<p class="jove_content">著者らは、手術法の支援のためにAdrianeモズレーとLibuse Jerabek（スタンフォード）に感謝したいと思います。 </p>

<p class="jove_content">すべての動物実験は、実験動物の管理と使用に関するUCLA​​の動物管理使用委員会のガイドラインと健康ガイドの国立研究所に従って行った。以下の手順の持続時間は、最初から最後まで約45〜60分である。 </p><p class="jove_title"> 1。手術野の調製</p><ol><li>きれいな動物の手術室での手順を実行します。 </li><li>設備：イソフルラン気化、加熱パッド付きGaymer Tポンプ。 </li><li>無菌ツール：2湾曲したピンセット、細いハサミ、針ホルダ。 </li><li>滅菌手袋は全体の手順の間に使用されなければならない。 </li></ol><p class="jove_title"> 2。動物の作製</p><ol><li>同じケージに同じ遺伝的背景から、同じような重さと大きさで、2雄または雌のマウスを配置し、調和のとれた共存を保証するために、少なくとも2週間を監視。雌マウスは、それらのより少なく積極的な行動のために好ましい。 </li><li>することにより、動物を麻酔イソフルラン気化器を使用する。ポジ·シール誘導チャンバ内でマウスをイソフルラン気化器（4-5％のV / V）に接続されている。麻酔が誘発されると、毛皮のシェービングエリアに動物を転送し、イソフルラン（1.5から2パーセントのV / V）に接続されているノーズコーンを通して作業中は麻酔を維持。ドライアイを防ぐために、Q-ティップで眼軟膏を適用します。 </li><li>仰臥位で動物を置きます。完全に右が少なくとも約1cmの1センチメートル膝下肘の上に載置された出発マウスの左右側に配置されたマウスの左側を剃る。 </li><li>無菌的に徹底的にアルコール綿続いベタジンに浸したワイプで（2〜3倍）を拭いて剃毛エリアを用意。無菌パッドでカバー加熱パッドの上にマウスを置きます。 </li><li>鎮痛のため、それぞれ10 mg / kgの0.1ミリグラム/ kgの用量で腹腔内または皮下にカルプロフェンおよびブプレノルフィンを管理します。 </li><li>で、背中合わせに、彼らの側に動物を配置上に向け剃毛分野をdjacent。手術領域の汚染を避けるため、操作領域を露出無菌ドレープでマウスを覆う。手術を行う際に、滅菌滞在する小さなドレープ開口部を作成します。私たちは、ビデオ撮影時に見やすくを持つようにドレープウィンドウを大きく。 </li></ol><p class="jove_title"> 3。並体結合</p><ol><li>鋭いはさみを使用して、肘0.5cm上で膝関節下の0.5センチメートル（ <strong>図1）</strong>へのすべての道を開始し、各動物の剃毛側面に縦方向の皮膚切開を行う。切開後、緩やかに湾曲鉗子で皮膚をかざして、皮​​下筋膜から皮膚をデタッチし、自由な皮膚の0.5cmに作成するために二対の筋膜を分離する。全体の切開に沿って、この分離を行う。 </li><li>他の右肘に1動物の左肘を取り付けることにより、接合を開始します。 olecranonsと膝関節の両方が目以下のはっきりと区別することができるEの皮膚切開。入社を容易にするために、最初のマウスの肘を曲げ肘頭下の非吸収性3-0縫合糸の針を渡します。同様に、第二のマウスの肘を曲げ、その下に、同じ縫合糸を通す。ダブル外科結びでしっかりとジョイントを取り付けます。 </li><li>同様の手順で膝関節を接続します。 </li><li>ジョイントの取り付けに続いて、ひざに向かって肘から腹始まる連続吸収5-0ビクリル縫合糸により2動物の皮膚に接続します。皮膚の破裂や剥離を防止する肘と膝の周りの領域における皮膚のタイトな縫合閉鎖を行う。腹側皮膚添付ファイルが完了すると、二重の手術結び目を実行します。腹臥位にマウスを置いて、二重の外科結びで終わる縫合背を続ける。縫合糸の連続性を確認して、開口部がないことを確認する。 </li><li> dehyを防ぐために、各マウスの皮下に0.9％NaClを0.5ミリリットルの管理dration。 </li></ol><p class="jove_title"> 4。術後の回復</p><ol><li>回復するまで加熱されたパッドの上に動物を保管してください。 </li><li>回復後、鎮痛剤のカルプロフェンとブプレノルフィンを提供しています。 （ステップ2.5）は、上記と同じ投与量で48時間、それぞれ、腹腔内または皮下に、24および12時間後に繰り返します。このような揺れなどの痛みや苦痛の徴候のために動物を監視し、尾の無気力、咀嚼は、 <em>など</em>グルーミングの欠如は、2週間毎日、バックアーチ。 </li><li>予防的に、細菌感染を防ぐために、10日間ミリリットル/トリムミリグラム2mgのサルファ剤/ mlの0.4彼らのウォーターボトルにスルファメトキサゾール/トリメトプリム経口懸濁液をマウスに扱う。 </li><li>家の寝具材料の吸引を防ぐために、モノリシックの寝具材料（ <em>例えば</em> 、紙タオルや吸収性の滅菌パッド）できれいにケージの各並体結合ペア。彼らは彼らの頭部を上にして胸骨横臥位を維持することができるとき、寝具いっぱいケージに動物を返します。歪みを最小限に抑えるために、並体結合の存在に調整しながら食品に手を伸ばすの、ケージの床に湿らせた食物ペレットを配置します。ネスティング材料を提供する。 1〜2週間で並体結合マウスは、外科的に対になっ前後と後肢で正常に歩き回るする能力を持っている。 </li><li>血液キメラ現象は、手術後10〜14日に行われます。 </li></ol><p class="jove_title"> 5。確認と逆の手順</p><ol><li> 2週間の血液キメラ現象（ <strong>図3）を</strong>確認するために、フローサイトメトリー分析のために各マウスの尾静脈から血液を引き出す後</li><li>フローサイトメトリー分析のための血液処理：10 mMのEDTAを500μlの血液を2〜3滴を希釈。 （PBSで希釈）の2％デキストランの500μl加え、完全に混合し、30分間37℃でインキュベートする。 、新しいチューブに上清を移し、5分間1,200 rpmでスピンし、フローサイトメトリー解析のために、PBS中のペレット化した細胞（または他のバッファ）懸濁します。 </li><li>実験設計に応じて、並体結合対は分離することができる後の時点でトン。我々の経験では、合併症のない最長9ヶ月間parabiosedペアを維持している。 </li><li>無菌外科表面に逆の手順を実行します。イソフルラン気化器（4-5％）に接続された導入室に配置することによってマウスを麻酔し（ステップ2.2）上記のように1.5〜2％イソフルランで麻酔を維持する。 </li><li> （2.3、2.4ステップ）初期縫合糸を周囲から髪を削除して、無菌的に上記のように剃らエリアを用意。 </li><li>鎮痛のために、それぞれ、10 mg / kgの0.1ミリグラム/ kgの用量で腹腔内または皮下カルプロフェン及びブプレノルフィンを投与する。 </li><li> （ステップ2.6）、上記のように、滅菌ドレープと手術領域をカバーしています。 </li><li>鋭いハサミを使用して、横方向の縫合糸に沿って縦切開を通してマウスを分離し、緩やかに湾曲鉗子のペアで2マウスとの間、新たに形成された筋膜を切り離す。関節を分離するために、縫合糸の結び目をカットそれらを接続する。滑らかなエッジを実現し、吸収性、連続5-0コーティングされたビクリル縫合糸で皮膚を再接続するために切開に沿って皮を切り落とす。 </li><li>脱水症を防ぐために、各マウスに0.9％のNaCl、皮下0.5mlのを投与。回復するまで加熱されたパッドの上に動物を保管してください。 </li><li>回復後、鎮痛剤を投与することを繰り返して、カルプロフェン（10 mg / kg）を24時間毎とブプレノルフィン（0.1 mg / kg）を48時間ごとに12時間。 </li><li>細菌感染を防止するために10日間、彼らのウォーターボトルに（2mgのサルファ剤/ mlの0.4 mg / mlのトリム）スルファメトキサゾール/トリメトプリム経口懸濁液をマウスに扱う。 </li></ol>

<ol>
	<li>Bert, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exp&#233;riences+et+consid&#233;rations+sur+la+greffe+animale.">Exp&#233;riences et consid&#233;rations sur la greffe animale.</a> <em style='font-style: italic'>Journal de l'Anatomie et de la Physiologie</em>. <b>1</b>, 69-87 .</li><li>Bunster, E., Meyer, R. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improved+methods+of+parabiosis.">Improved methods of parabiosis.</a> <em style='font-style: italic'>Anat. Rec</em>. <b>57</b>, 339-380 (1933).</li><li>Waskow, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generation+of+parabiotic+mice+for+the+study+of+DC+and+DC+precursor+circulation.">Generation of parabiotic mice for the study of DC and DC precursor circulation.</a> <em style='font-style: italic'>Methods Mol. Biol</em>. <b>595</b>, 413-428 (2010).</li><li>Finerty, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Parabiosis+in+physiological+studies.">Parabiosis in physiological studies.</a> <em style='font-style: italic'>Physiol. Rev</em>. <b>32</b>, 277-302 (1952).</li><li>Aicher, A., Heeschen, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nonbone+marrow-derived+endothelial+progenitor+cells:+what+is+their+exact+location.">Nonbone marrow-derived endothelial progenitor cells: what is their exact location.</a> <em style='font-style: italic'>Circ. Res</em>. <b>101</b>, e102 (2007).</li><li>Abe, S., Boyer, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cells+derived+from+the+circulation+contribute+to+the+repair+of+lung+injury.">Cells derived from the circulation contribute to the repair of lung injury.</a> <em style='font-style: italic'>Am. J. Respir. Crit. Care Med</em>. <b>170</b>, 1158-1163 (2004).</li><li>Donskoy, E., Goldschneider, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thymocytopoiesis+is+maintained+by+blood-borne+precursors+throughout+postnatal+life.+A+study+in+parabiotic+mice.">Thymocytopoiesis is maintained by blood-borne precursors throughout postnatal life. A study in parabiotic mice.</a> <em style='font-style: italic'>J. Immunol</em>. <b>148</b>, 1604-1612 (1992).</li><li>Powell, A. E., Anderson, E. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fusion+between+Intestinal+epithelial+cells+and+macrophages+in+a+cancer+context+results+in+nuclear+reprogramming.">Fusion between Intestinal epithelial cells and macrophages in a cancer context results in nuclear reprogramming.</a> <em style='font-style: italic'>Cancer Res</em>. <b>71</b>, 1497-1505 (2011).</li><li>Duyverman, A. M., Kohno, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+transient+parabiosis+skin+transplantation+model+in+mice.">A transient parabiosis skin transplantation model in mice.</a> <em style='font-style: italic'>Nat. Protoc</em>. <b>7</b>, 763-770 (2012).</li><li>Ajami, B., Bennett, J. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Local+self-renewal+can+sustain+CNS+microglia+maintenance+and+function+throughout+adult+life.">Local self-renewal can sustain CNS microglia maintenance and function throughout adult life.</a> <em style='font-style: italic'>Nat. Neurosci</em>. <b>10</b>, 1538-1543 (2007).</li><li>Weissman, I. L., Jerabek, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tolerance+and+the+H-Y+antigen:+Requirement+for+male+T+cells,+but+not+B+cells,+to+induce+tolerance+in+neonatal+female+mice.">Tolerance and the H-Y antigen: Requirement for male T cells, but not B cells, to induce tolerance in neonatal female mice.</a> <em style='font-style: italic'>Transplantation</em>. <b>37</b>, 3-6 (1984).</li></ol>

<p class="jove_content">著者らは、競合する金融利害関係はありません。 </p>

<p class="jove_content">ここで説明並体結合の方法は、低死亡率の最低限の技術的な問題と結果を示す。膝や肘の関節の取り付けはBunsterとマイヤー技術の大幅な改善である。しかし、手順はこのように、手術を通して無菌状態の維持が不可欠である侵襲性のままになります。さらに、手術部位の感染を防止するためには、parabiosed動物は抗生物質の組み合わせを受信し、定期的に監視することが重要である。しっか?...

<p class="jove_content">並体結合、2生物の外科的接合は、最初に共有循環系を研究するためのモデルを開発するための方法として、ポール·バートによって1864に記載され、2匹のラット<sup>1</sup>の皮膚や筋肉の壁の接合で構成された。並体結合は、炎症<sup>3</sup>部位での微小血管の形成を促進し、このような下垂体および生殖腺だけでなく、高血圧<sup>4</sup>における腎臓の役割の間のホルモン通信などの生理学的研究にはいくつかのアプリケーションを、持っていた。さらに、血管新生<sup>5、</sup>造血幹細胞<sup>6</sup>の移動、およびリンパ球輸送<sup>7、</sup>ならびに腫瘍転移<sup>8,9</sup>における炎症性または幹細胞を循環させる役割と動態に前駆細胞の動員および組込みを調べるために使用されているおよび神経変性疾患<sup>10。</sup> </p><p class="jove_content">並体結合の嘘の一つの重要な利点その中で提携した動物は、免疫学的反応を誘発することなく、細胞移動および血管新生を可能にする一般的な循環抗原を共有しています。重要なことには、ワイズマン<em>らは、</em>雄と雌のマウス間の並体結合は、抗HY抗体<sup>11</sup>の形成をもたらさない示した。 </p><p class="jove_content">ポール·バートによって記述、元のプロトコルでは、2匹の動物は、皮膚と筋肉の壁<sup>1</sup>の接続を介して接合した。しかしながら、この方法は、動物に重大な歪みを引き起こし、傷口の感染による高い死亡率となった。それ以来、並体結合技術が最も優勢が1933 <sup>2</sup>でBunsterとマイヤーによって提案されたプロトコルであることをいくつかのグループにより改訂されました。彼らの方法は、動物のためのより良いサポートが少なく、痛みを許可する、肩甲骨の関節、体腔と皮膚の接合含まれています。同時に、新しい方法は、最小限の術後ケアとsignifiの結果cantlyは死亡率を減少させた。本明細書に記載されたプロトコルは、低侵襲性であり、強固な接合が可能とBunsterマイヤー法の改良法である。すなわち、マウスは肘と膝関節、並びに皮膚を介して接続されている。したがって、これは皮膚のを防止拡張子を結合し、痛みが少なく、合併症を引き起こす。ここでは、マウスを表現する構成的GFPに野生型（WT）成体マウスの接合について説明します。私たちは、手術後2週間は、我々が共有循環系を作成するために、この外科処置の有効性を実証する血液キメラの50％を達成できることを示している。 </p>

<table><tbody><tr><td>Isoflurane-Phoenix</td><td>Clipper</td><td>NDC 57319-559-06</td><td></td></tr><tr><td>Posi-Seal Induction Chamber</td><td>Molecular Imaging Products</td><td>AS-01-0530-SM</td><td></td></tr><tr><td>Portable Anesthesia System</td><td>Molecular Imaging Products</td><td>AS-01-0007</td><td></td></tr><tr><td>Gaymer T Pump</td><td>Gaymar Industries, Inc</td><td>TP650</td><td></td></tr><tr><td>Warming Blanket (Heating pad)</td><td>Kent Scientific Corp</td><td>TP-22G</td><td></td></tr><tr><td>Curved forceps</td><td>Roboz</td><td>RS-5101</td><td></td></tr><tr><td>Scissors</td><td>Fine Science Tools (FST)</td><td>FST 14063-09</td><td></td></tr><tr><td>Needle holder</td><td>FST</td><td>FST 12501-13</td><td></td></tr><tr><td>Electrical shaver</td><td>Oster</td><td>Golden A5</td><td></td></tr></tbody></table>

parabiosis in mice: a detailed protocol

<p class="jove_content">並体結合は、血液循環の共有を可能にする2つの生物の外科的和集合である。 2動物の皮膚を取り付けることは、炎症部位での微小血管の形成を促進する。並体結合パートナーは、循環抗原を共有するため、有害な免疫反応の自由である。まず1864年<sup>1</sup>でポール·バートによって記述、並体結合手術は動物の生存<sup>2を</sup>改善するために1933年にBunsterとマイヤーによって精製した。現在のプロトコルでは、2匹のマウスを外科Bunsterとマイヤー法の改良法以下の接合されている。動物は縫合し、皮膚に負担を防ぐ強固なサポートを可能にする皮膚の取り付けに続いて、肘と膝関節を介して接続されている。ここで、我々は詳細に野生型（WT）マウスにマウスを表現するユビキタスGFPの合流並体結合を記述します。血液循環液の循環に二週間処置後、ペアを分離し、GFP陽性細胞をフローサイトメトリー分析によって検出することができるWTマウスのイオン。血液キメラ現象は、一方が他方にある1動物からの循環細胞の寄与を調べることができます。 </p>

<p class="jove_content"> 2の生物の並体結合の予想結果は、一般的な血液循環（ <strong>図2）</strong>に、それぞれの動物の循環系の等しい貢献である。一つは、簡単にフローサイトメトリー分析によってparabiosed WT及びGFP陽性マウスの血液の正常な平衡を確認することができます。ここでは、静脈血を、手術後2週間目に両方parabiontsの尾から得られ、（赤血球を除くために）、末梢血細胞を分画した。分画された造血由?...

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Parabiosis in Mice: A Detailed Protocol

<p class="jove_content"> 2つの生物の結合並体結合は、共有循環系の開発につながる。このプロトコルでは、野生型マウスと恒常的GFP発現マウスとの間の並体結合接続を形成するための手術ステップを記載している。 </p>

マウスの並体結合：詳細なプロトコル

Parabiosis is a surgical union of two organisms allowing sharing of the blood circulation. Attaching the skin of two animals promotes formation of ...

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Medicine

74.2K ビュー.  University of California, Los Angeles.  2つの生物の結合並体結合は、共有循環系の開発につながる。このプロトコルでは、野生型マウスと恒常的GFP発現マウスとの間の並体結合接続を形成するための手術ステップを記載している。 

ビデオ: Parabiosis in Mice: A Detailed Protocol

Genetics of Organisms

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">REF and WEF Fly Genetics
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">NOTE: At your station, you should have four vials of flies. They will be labeled as male or female. They will also be labeled with the notation REF for red-eyed flies or WEF for white-eyed flies. The female vial should also be labeled virgin as these flies should be unmated for the purpose of this experiment.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Make sure you have around five flies in each vial. NOTE: You will perform crosses using these flies to determine if red or white eyes is dominant and if the trait is sex-linked. You should also have two fly anesthesia wands, a paint brush, magnifying glass, fly anesthetic, some empty fly vials, and a sheet of white paper.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Draw a line in the middle of the paper to make a male side and a female side.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Dip the fly anesthesia wand into the solution and insert it into the red-eyed female vial until the flies stop moving.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Then, dip the same wand into the white-eyed male vial until these flies are also anesthetized.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">Tap the flies from the two vials onto the sheet - Females on one side and males on the other.
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Check to make sure that all the flies from the female vial are female and likewise, confirm that the male flies are male.  NOTE: Males are slightly smaller and have a darker solid pigment on the end of their abdomen. Females are larger and have stripes rather than a solid spot.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">If there are any male flies in the female vial, obtain a new vial as these females likely have sperm from males from their own strain and are unusable.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">Now, move the five red-eyed females and five white-eyed males into a new vial using the paintbrush.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">Label this vial to indicate that this is a cross between red-eyed females and white-eyed males. HYPOTHESES: Use Punnett squares to formulate hypotheses for this experiment. Consider what you will see in a few days if the white allele is dominant and there is no effect of parental sex. What if red is dominant? Under the potential offspring genotype in each box, write the expected phenotype.
</div></li><li data-sub-note-item="1-11"><div class="lab-step">To set up the reverse cross, again dip the fly anesthesia wand into the solution and insert it this time into the white-eyed female vial and then the red-eyed male vial until all the flies have stopped moving.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">Tap the flies onto a sheet of white paper and check to make sure that all the flies from the female vial are female and likewise with the male flies.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">Now, move the five white-eyed females and five red-eyed males into a new vial using the paintbrush.
</div></li><li data-sub-note-item="1-14"><div class="lab-step">Label this vial to indicate that this is a cross between white-eyed females and red-eyed males.
</div></li><li data-sub-note-item="1-15"><div class="lab-step">Finally, place the two tubes containing the new crosses into a rack for incubation at room temperature. NOTE: After about three days, larvae should be moving around in the vial.
</div></li><li data-sub-note-item="1-16"><div class="lab-step">Remove the 10 adult flies and dispose of them into the morgue tube containing 70% ethanol.
</div></li><li data-sub-note-item="1-17"><div class="lab-step">Now, put the tubes back in the rack and let the larvae mature until they reach the adult stage in up to two weeks.
</div></li><li data-sub-note-item="1-18"><div class="lab-step">Once all of the larvae from these crosses have matured to adulthood, anesthetize the flies in one of the tubes using the wand.
</div></li><li data-sub-note-item="1-19"><div class="lab-step">Then, on a piece of white paper, sort the flies by sex and phenotype and then record these data.
</div></li><li data-sub-note-item="1-20"><div class="lab-step">Now, repeat this process and tally the sexes and phenotypes for the second cross (steps 1 &#8211; 18).
</div></li><li data-sub-note-item="1-21"><div class="lab-step">Finally, once you have counted all of your flies, dispose of them in the morgue vial.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">To begin investigating your results, first compare the data you collected with the hypothesis you made using the Punnett squares.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Did your data match the simple Punnett square hypothesis where red or white is dominant and there is no effect of sex? Compare your data with other lab groups.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">If your data does not match what your Punnett squares predicted, does this indicate that the gene may be sex-linked?
</div></li><li data-sub-note-item="2-4"><div class="lab-step">If the red gene were dominant and sex-linked, when we cross red-eyed females with white-eyed males, all of the offspring would have red eyes. Assuming all of the females were homozygous for the color allele, does this match your data for the first cross?
</div></li><li data-sub-note-item="2-5"><div class="lab-step">If white were dominant and the gene were sex-linked, we would see the exact opposite. So all of the flies would have white eyes. From your data, can you conclude from your first cross whether red or white is dominant?
</div></li><li data-sub-note-item="2-6"><div class="lab-step">When the reciprocal cross of white-eyed females and red-eyed males is performed, if the red phenotype is dominant, all of the females would be red and all of the males would be white. This would also confirm that the gene is specifically x-linked because white-eyed females could then only be produced in crosses with white-eyed males. Did your second cross produce any white-eyed females?
</div></li><li data-sub-note-item="2-7"><div class="lab-step">Finally, if the gene were x-linked and white was dominant, crossing white-eyed females with red-eyed males would result in all of the offspring having white eyes. Does this hypothesis fit with your second cross?
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Look over your data and conclude which of the hypotheses best fits the patterns you found in your crosses. If your data does not fit with any of these, hypothesize why this might be the case.
</div></li></ul></li></ol>

Genetics of Organisms: REF and WEF Fly Genetics - Procedure

生物の遺伝学

REF and WEF Fly Genetics
ExpandNOTE: At your station, you should have four vials of flies. They will be labeled as male or female. They will also be ...

Education

JoVE Lab Manual

生物学

遺伝学

Lab Manual Sub Articles

Bacterial Transformation

<ol class="note-items"><li data-note-item="1" data-parent-collapse=""><strong>Bacterial Transformation</strong>
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">IMPORTANT: In addition to wearing the appropriate personal protective equipment, be sure to take care to keep your face away from suspension cultures, and to avoid inhaling reagents. Do not touch your face while performing the experiment. Always wash your hands before and after every experiment.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">When you are ready to safely begin, check to make sure you have a tube or tubes of competent <em>E. coli</em> cells in your ice bucket.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Then, transfer 50 &#181;L from one of these tubes to each of two new micro-centrifuge tubes, one labeled '&#8211;', and one labeled '+'.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Next, pipet 2 &#181;L of p-GREEN plasmid into the plus tube and flick the tube gently to incorporate the plasmid throughout the bacterial solution. HYPOTHESES: The experimental hypothesis is that on plates containing no antibiotic, bacteria with or without the plasmid can survive, and so both will grow lawns of bacteria. Conversely, on the plates containing the ampicillin antibiotic, only the bacteria successfully transformed with the resistance plasmid will grow, whereas the minus plasmid plates will contain no bacteria. The null hypothesis of this experiment is that there will be no difference in bacterial growth between the plates.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Incubate both tubes on ice.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">After 30 minutes submerge both tubes 3/4 of the way into the water of a 42 &#176;C water bath for 30 seconds.
</div></li><li data-sub-note-item="1-7"><div class="lab-step">At the end of the water bath incubation, place the tubes back on ice for 5minutes, then add 950 &#956;L of room temperature SOC media to each tube.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">Incubate the tubes in a shaker at 37 &#176;C for 60 minutes at 250 rpm.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">While the tubes are incubating, label the bottoms of one LB and one LB/ampicillin, or AMP plate, &#8220;+ plasmid&#8221;, and the bottoms of one LB plate and one LB/AMP plate &#8220;- plasmid&#8221;.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">At the end of the incubation, pipet 50 &#956;L from the &#8220;+ plasmid&#8221; cell tube onto both the LB/AMP plus plate and the LB plus plate.
</div></li><li data-sub-note-item="1-11"><div class="lab-step">Then, with a fresh tip, add 50 &#956;L of the bacteria that were not exposed to the plasmid to each of the minus plates.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">Then, spread each plate using a fresh pipette tip.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">Cover the plates with their lids, and then seal them with laboratory film.
</div></li><li data-sub-note-item="1-14"><div class="lab-step">Then store the plates upside down in a 37 &#176;C incubator for 24 hours.
</div></li></ul></li><li data-note-item="2" data-parent-collapse=""><strong>Results</strong>
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">The day after the transformation, observe which plates have bacteria, and how the bacteria are distributed over each plate.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Record the number of colonies and the color of the colonies for different conditions. <a href="https://www.jove.com/CDNSource/lm/labs/11/Table_1_Bacterial_Transformation.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="2-3"><div class="lab-step">To determine the initial mass, IM, of the plasmid, first multiply the plasmid concentration from the stock tube of p-GREEN by the volume transferred into the experimental tube.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">To calculate how much DNA is spread onto each plate, divide the volume of bacterial suspension spread onto each plate by the volume of the suspension in the tube, and multiply this fraction by the initial mass of plasmid used.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">To determine the transformation efficiency, count all of the visibly distinct colonies on the plus plasmid LB/AMP plate, and divide this number by the mass of plasmid spread. NOTE: The unit for transformation efficiencies is cfu/&#956;g of plasmid DNA.
</div></li><li data-sub-note-item="2-6"><div class="lab-step">Record all of the results for each plate in the table and analyze them.
</div></li><li data-sub-note-item="2-7"><div class="lab-step">Observe any differences between the colonies and make hypotheses to explain this observation. Consider the factors that may affect the transformation efficiency of competent bacteria.
</div></li><li data-sub-note-item="2-8"><div class="lab-step">NOTE: You will have also noticed that each plate had a different outcome. The minus plasmid and plus plasmid LB plates should have grown a lot of bacteria across their plates. The minus plasmid LB/AMP plate should have no bacterial growth. And the plus plasmid LB/AMP plate should have grown bacterial colonies that fluoresced green.
</div></li></ul></li></ol>

Bacterial Transformation using Plasmids - Procedure

細菌の形質転換

Bacterial Transformation
ExpandIMPORTANT: In addition to wearing the appropriate personal protective equipment, be sure to take care to keep your face ...

Group Behavior

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">The Hawk-Dove Game
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">NOTE: In this activity, you will play the hawk and dove game with a single partner and then with multiple partners to observe hypothetical costs and benefits of the antagonist or cooperative behavior respectively. 
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Before the activity starts, review the cost-benefit table so that you understand how to score the game. <a href="https://www.jove.com/CDNSource/lm/labs/33/Table_1_Group_behavior.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="1-3"><div class="lab-step">HYPOTHESES: The experimental hypothesis for this exercise could be that the dove strategy will become more frequent between repeated partners, and the hawk strategy will become more frequent when interactions are with a variety of partners. The null hypothesis could be that there will be no change in strategy over time with repeat or different partner combinations. NOTE: Benefit can be determined by looking at the left column and choosing the row that corresponds to the card you played. The box that corresponds to the choice of your partner can be found in the top row. For the purposes of this game, B equals four and C equals three. So depending on the cards played, player one here, which is you, would gain a score of two, four, zero or one half. 
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Collect a set of two game cards. 
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Take out the cards labeled hawk and dove and place them face down on the table or conceal them in your hands. 
</div></li><li data-sub-note-item="1-6"><div class="lab-step">When the instructor gives the signal show either of the two cards to the student sitting across from you. Try to strategize how to receive the highest benefit from every interaction on your own but do not communicate with your partner. 
</div></li><li data-sub-note-item="1-7"><div class="lab-step">In the Single Partner Recording Table (Table 2) provided by your instructor, note the cards played as well as the benefit you received for that round.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">Repeat showing cards to the same partner and determining benefit on your instructor&#39;s signal for a total of 15 rounds.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">At the end of those rounds, the student with the highest total benefit value was determined to be the &#8220;winner&#8221;, or the most fit.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">As a class, determine the percentages of hawks and doves played as well as the average benefit received for each round, using this formula: Class average benefit=(sum total benefit)/(number of players)
</div></li><li data-sub-note-item="1-11"><div class="lab-step">For the second game, and then for each round of play, one row of students should rotate to their right, leaving all players with a new partner.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">Continue playing for 15 rounds or until students end up back in their starting positions, recording the data in the Multiple Partner Recording Table (Table 3) given to you by your instructor.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">For each round, determine the percentages of hawks and doves played that round, as well as the average benefit received.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Cheating and Cooperation Exercise
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">NOTE: In this activity, you will exchange benefits, represented by colored or similar items, with your classmates. In this simulation, a cheater is defined as a person who gives fewer benefits than they receive. During this activity, you will observe the frequency of cheating behavior over time. 
</div></li><li data-sub-note-item="2-2"><div class="lab-step">HYPOTHESES: The experimental hypothesis is that cheaters will receive fewer benefits over time and cooperators will continue to provide equal benefits to one another. The null hypothesis is that the frequency of cheaters and cooperators does not change over time. 
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Each person should collect beads of one color to represent a tangible benefit, as well as one paper bag and a three-ring binder.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Get into a group of three so that all members have objects of the same color.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Now find another group with a different color to pair with. In each round, partners will exchange benefits without knowing what they are receiving from their partner and without discussion within their group.  NOTE: Benefits of a different color from your starting color are worth twice as much.
</div></li><li data-sub-note-item="2-6"><div class="lab-step">Turn face to face with a member from the other group while still being able to conceal the objects in your possession.  TIP:<strong> </strong>Setting up a screen using a notebook or sitting behind a desk can be helpful here.
</div></li><li data-sub-note-item="2-7"><div class="lab-step">When given a signal from the instructor, place zero to two benefits into the paper bag and pass it to your partner.
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Simultaneously, receive the bag from your partner and then remove the benefits from the bag.
</div></li><li data-sub-note-item="2-9"><div class="lab-step">Make a note of how many benefits you gave away and how many you received in the Cheating and Cooperation Table (Table 4) provided by your instructor.
</div></li><li data-sub-note-item="2-10"><div class="lab-step">Turn to a new member from the other group and repeat the exchange.
</div></li><li data-sub-note-item="2-11"><div class="lab-step">Rotate through all of the members of the other group for as many rounds as the instructor determines - which will not be revealed until the game ends so as not to alter player behavior.
</div></li><li data-sub-note-item="2-12"><div class="lab-step">Once the game has ended, note for each round in the cheating and cooperation table whether you gave more than your partner, received more from them or if you gave the same amount.
</div></li><li data-sub-note-item="2-13"><div class="lab-step">Now total up all of the benefits in your possession, with benefits you&#39;ve received from the other group being worth twice as much.
</div></li><li data-sub-note-item="2-14"><div class="lab-step">For each round, the whole class should total up the number of cheaters - with cheaters defined as those who gave less than they received.
</div></li></ul></li><li data-note-item="3" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="3-1"><div class="lab-step">For both the single partner hawk-dove game and the multiple partners hawk-dove game, use the whole class data to plot the percentages as two separate lines - one for the percentage of hawks, and one for the percentage of doves, for each of the 15 rounds.
</div></li><li data-sub-note-item="3-2"><div class="lab-step">Take note of whether the cards played changed as more rounds were played.
</div></li><li data-sub-note-item="3-3"><div class="lab-step">Manipulate the values for B and C to see if you can find the point at which values of B and C would render the hawk strategy no longer effective (this would be the point when there is no benefit to choosing the hawk strategy over always choosing the dove strategy).
</div></li><li data-sub-note-item="3-4"><div class="lab-step">As a class, use the data from the cheating and cooperation activity to make a graph showing the percentage of rounds each individual player cheated versus the amount of benefits they ended with.
</div></li><li data-sub-note-item="3-5"><div class="lab-step">Consider whether it is better to be a cheater or a cooperator in this scenario and discuss as a class what makes this scenario different from the hawk-dove game.
</div></li><li data-sub-note-item="3-6"><div class="lab-step">Now, use the class data to graph the percentage of cheaters against the number of rounds, and look for any trends over time.
</div></li><li data-sub-note-item="3-7"><div class="lab-step">Consider which type of strategy would most likely evolve in a setting where partners interact repeatedly.
</div></li><li data-sub-note-item="3-8"><div class="lab-step">Then, note which tactic might be beneficial for a species that relies on cooperation. 
</div></li><li data-sub-note-item="3-9"><div class="lab-step">Discuss as a class what would likely happen if you increased or decreased the relative value of the benefits you receive.
</div></li></ul></li></ol>

Group Behavior: Hawk-Dove Game and Trading Exercise - Procedure

グループの動作

The Hawk-Dove Game
ExpandNOTE: In this activity, you will play the hawk and dove game with a single partner and then with multiple partners to observe ...

生態学

Optimal Foraging

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Simulating Foraging with Artificial Habitats and Prey
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">NOTE: The foragers will hunt for prey represented by pinto beans in four buckets of rice with varying prey densities. Without knowing what these densities are, foragers must obtain as many prey items as possible in as little time as possible. 
</div></li><li data-sub-note-item="1-2"><div class="lab-step">HYPOTHESES: In this experiment, the experimental hypothesis could be that foragers will catch the most prey in the higher prey density bucket and also spend the most time foraging there.  The null hypothesis would then be that foragers will catch an equal number of prey in each bucket and that foragers will spend an equal amount of time foraging in each patch. 
</div></li><li data-sub-note-item="1-3"><div class="lab-step">To begin the foraging activity, record the names and role of each participant at the top of the student data collection sheet.  NOTE: Before starting the experiment, the forager is allowed to become familiar with the foraging process &#8211; the following steps are a practice round. 
</div></li><li data-sub-note-item="1-4"><div class="lab-step">The forager should kneel or sit in front of the average density bucket. 
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Once the timer says start, the forager may begin looking for the pinto beans representing the prey. The forager can spend as much time as they like foraging in this bucket. 
</div></li><li data-sub-note-item="1-6"><div class="lab-step">Once a prey item is located, the forager must signal that one has been found and place the prey in the cup next to the bucket.
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Once they believe they have acquired most of the prey in the patch, they should tell the timer to stop the time and the practice round is over. 
</div></li><li data-sub-note-item="1-8"><div class="lab-step">To begin the simulation proper, the forager should sit or kneel in front of bucket A. 
</div></li><li data-sub-note-item="1-9"><div class="lab-step">The timer must be ready to start the stopwatch. 
</div></li><li data-sub-note-item="1-10"><div class="lab-step">The recorder should be ready to record the time each prey item is located, in addition to the time that the forager arrived at and left each bucket. 
</div></li><li data-sub-note-item="1-11"><div class="lab-step">When all parties are ready, the timer should start the stopwatch and say &#8220;start&#8221;,  and the forager should begin foraging. NOTE: The stopwatch will run through the rest of the round for buckets B and C.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">As the forager signals finding pinto beans and places them in the cup next to the bucket, the timer should tell the recorder the time.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">The recorder should then write the time in the student data collection sheet.  <a href="https://www.jove.com/CDNSource/lm/labs/34/Table_1_Optimal_foraging.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="1-14"><div class="lab-step">Once the forager has decided to move on to the next bucket, they should tell the timer &#8220;done&#8221;, and begin moving to the next patch. 
</div></li><li data-sub-note-item="1-15"><div class="lab-step">The recorder should record the time the forager left and the time they arrived at bucket B in the student data collection sheet.
</div></li><li data-sub-note-item="1-16"><div class="lab-step">Repeat the same procedure for buckets B and C. Once the forager has completed the activity, it is time for the next forager to begin using the same procedure. 
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">Before performing any data analysis, review the data collection sheet and compile your individual data for captured prey, foraging time, and travel time for each bucket in your student calculations table. <a href="https://www.jove.com/CDNSource/lm/labs/34/Table_2_Optimal_foraging.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 2</a>
</div></li><li data-sub-note-item="2-2"><div class="lab-step">After this, calculate your individual capture rate for each bucket by dividing the number of prey captured by the sum of the foraging time and travel time and record these values in your table. Individual capture rate =  Number of prey captured / (foraging time + travel time)
</div></li><li data-sub-note-item="2-3"><div class="lab-step">To calculate your Giving-Up-Time or a GUT for each bucket, subtract the time that the last prey item was acquired from the time you left the bucket.  GUT = time forager left bucket  -  time last prey item acquired
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Next, calculate the mean GUT for your foraging group and record the values in your table.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Finally, calculate your scaled GUT for each bucket by dividing the bucket GUT by your group&#39;s mean GUT. Scaled GUT =  Bucket GUT / Group mean GUT NOTE: The scaled GUT will allow class data to be pooled later on without differences in individual foragers skewing the data. 
</div></li><li data-sub-note-item="2-6"><div class="lab-step">The instructor should now compile the data into the classroom spreadsheet.  <a href="https://www.jove.com/CDNSource/lm/labs/34/Table_3_Optimal_foraging.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 3</a>
</div></li><li data-sub-note-item="2-7"><div class="lab-step">Plot a bar graph of the average number of prey collected in each bucket. On the x-axis, A should represent the low prey density bucket, B should represent the medium prey density bucket, and C the high prey density bucket. Plot the y-axis as the number of prey items caught. 
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Next, plot the graph of the class average scaled GUT, where the x-axis represents the buckets with varying levels of prey density and the y-axis shows the values for the average scaled GUT. 
</div></li><li data-sub-note-item="2-9"><div class="lab-step">Plot a third graph of the average time spent foraging in each bucket. 
</div></li><li data-sub-note-item="2-10"><div class="lab-step">Now review the Marginal Value Theorem (MVT) from the concepts video and examine the three graphs that you generated from your class data. Take note of whether the graphs conform to the predictions of MVT. 
</div></li></ul></li></ol>

Optimal Foraging: Foraging with Artificial Habitats and Prey - Procedure

最適な採餌

Simulating Foraging with Artificial Habitats and Prey
ExpandNOTE: The foragers will hunt for prey represented by pinto beans in four buckets of rice with ...

Sexual Selection and Mate Choice

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Sexual Selection Class Simulation
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">Note: In this activity, you and your classmates will portray a sexually reproducing population consisting of an equal number of male and female animals. You will simulate several different mating scenarios to determine how relative quality, attractiveness, sex, and parental commitment affect mate choice and reproductive success. The goal of this activity is to maximize your progeny. 
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Hypotheses: There are several overall hypotheses for female and male behavior for this activity. The main experimental hypothesis for female behavior is that females will mate with males of high quality more frequently than those of low quality, yielding a higher reproductive success. The null hypothesis is that females will not demonstrate preference towards high- or low-quality males in any of the four rounds of mating. For males, one experimental hypothesis is that males with a higher visible score will have greater reproductive success. Another could be that males will be more selective for females in scenarios in which they join females to care for offspring as opposed to scenarios in which they exhibit no parental commitment. The null hypothesis for male behavior might be that males will not demonstrate preference towards high- or low-quality females in any of the four rounds of mating. 
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Activity Overview: To begin, each student will be assigned a sex as well as a relative quality and attractiveness score. Remember - the goal is to maximize your reproductive success in each round. You do this by mating with partners of the highest possible quality which is rated on a scale of one to three with three being the highest possible score. Attractiveness scores, which follow the previously described scoring paradigm, correlate with quality but do not exactly predict the score. Though useful, they are not a perfect way to choose the best mate.  If you are assigned a male ID holder, you will have 16 gametes. However, if you are assigned a female ID holder, you will only have four. A successful mating occurs when one sperm and one egg are paper-clipped together and deposited at the fertilization area. 
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Note - Rules of the activity: 
<ul>
	<li>Assigned male and female identities for these experiments are not representative of students&#39; personal gender identities. </li>
	<li>Running is not permitted. </li>
	<li>Talking is not permitted.</li>
	<li>Hidden values cannot be shared but individuals can attempt to hide their visible scores. </li>
	<li>Respect the space around each individual and remain an arm&#39;s length away. </li>
	<li>Offspring will only be counted towards an individual score if both gametes are paper-clipped together and dropped off in the designated fertilization area before the end of the two-minute round. </li>
</ul>
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Round 1 &#8211; Hidden Quality Score
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">To begin the first simulation, take the clip-on ID holder you were randomly assigned and make sure it contains four gametes if you are assigned female or sixteen gametes if male. Males should also have a strip of white paper and two paperclips. 
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Next, males should paperclip the white strip of paper over their quality score to hide it, but keep the attractiveness score visible. 
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Record your ID number for round one in table two.  <a href="https://www.jove.com/CDNSource/lm/labs/35/Table_2_sexual_selection.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 2</a>
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Next, clip on the ID holder and move to the designated mating area. 
</div></li><li data-sub-note-item="2-5"><div class="lab-step">When the instructor says begin, you may commence the mating simulation. You will have exactly two minutes to produce as many offspring as possible. Males may freely give their sperm to females, and females may choose to use these sperm immediately after which they would walk to the fertilization area and paperclip it to one of their eggs, or they may choose to wait until they receive other sperm to do so. 
</div></li><li data-sub-note-item="2-6"><div class="lab-step">Once the gametes have been deposited in the fertilization area, females may resume mating. 
</div></li><li data-sub-note-item="2-7"><div class="lab-step">When the two minutes have elapsed, all matings must stop and any gametes not in the fertilization area at this time will not be counted. 
</div></li><li data-sub-note-item="2-8"><div class="lab-step">The instructor should now tally all of the matings and record the numbers of the individuals who produced each offspring.
</div></li><li data-sub-note-item="2-9"><div class="lab-step">Now, find all the gametes that came with your ID holder and repackage them for the next round. 
</div></li></ul></li><li data-note-item="3" data-parent-collapse="">Round 2 &#8211; Hidden Attractiveness Score
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="3-1"><div class="lab-step">For the second round, obtain a new randomly assigned ID holder and again ensure it contains the proper number of gametes. 
</div></li><li data-sub-note-item="3-2"><div class="lab-step">Next, males should use the paperclip and strip of white paper to hide their attractiveness score (instead of the quality score, which was covered in round one).
</div></li><li data-sub-note-item="3-3"><div class="lab-step">Record the ID number of your clip in table two under round two.  Note: The mating simulation process for round two will be the same as round one.
</div></li><li data-sub-note-item="3-4"><div class="lab-step">Move to the designated mating area and with for the instructor to signal the start of the round. As before, males may freely give away their sperm while females will choose to use it now or wait for different sperm. When a female decides to fertilize an egg, she will walk to the fertilization area and paperclip the gametes together. 
</div></li><li data-sub-note-item="3-5"><div class="lab-step">Proceed with the simulation for two minutes, and at the signal from the instructor all mating activity should stop. 
</div></li><li data-sub-note-item="3-6"><div class="lab-step">The instructor should now tally all of the matings from this second round.
</div></li><li data-sub-note-item="3-7"><div class="lab-step">Now, find and return the gametes to your ID holder, and then return the holder to the instructor.
</div></li></ul></li><li data-note-item="4" data-parent-collapse="">Round 3 &#8211; Visible Quality and Attractiveness Scores
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="4-1"><div class="lab-step">For the third simulation, you should again obtain a new clip-on ID holder and check that it contains the appropriate number of gametes. Record the ID number of your clip in table two.
</div></li><li data-sub-note-item="4-2"><div class="lab-step">For this round, leave both attractiveness and quality scores visible on male IDs. Note: The mating simulation process for round three will be the same as the previous two rounds.
</div></li><li data-sub-note-item="4-3"><div class="lab-step">Move to the mating area and wait for the signal to begin the simulation.
</div></li><li data-sub-note-item="4-4"><div class="lab-step">Once the instructor starts the timer and gives the signal, commence mating. As before, males are allowed to freely give away their sperm to females and females may use the sperm immediately or wait for different sperm. Again, fertilized eggs only count if placed in the fertilization area. 
</div></li><li data-sub-note-item="4-5"><div class="lab-step">After the two-minute time limit, the instructor should tally all matings. 
</div></li><li data-sub-note-item="4-6"><div class="lab-step">Now, find and return the gametes to your ID holder, and then return the holder to the instructor.
</div></li></ul></li><li data-note-item="5" data-parent-collapse="">Round 4 &#8211; Increased Male Parental Care
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="5-1"><div class="lab-step">In the final round, males will again leave their attractiveness and quality scores visible. 
</div></li><li data-sub-note-item="5-2"><div class="lab-step">Obtain an ID holder and ensure it contains the appropriate number of gametes. 
</div></li><li data-sub-note-item="5-3"><div class="lab-step">Clip on the ID holder and move to the mating area. Note: As before, males may freely give away their sperm to females. However, in this round, once a female chooses to mate with a male, both the male and female will walk to the fertilization area to paperclip their gametes together. After placing the paper-clipped gametes on the table, both male and female actors may return to the mating area. 
</div></li><li data-sub-note-item="5-4"><div class="lab-step">The instructor should signal the start of the round and begin the timer &#8211; at this point participants can begin the simulation.
</div></li><li data-sub-note-item="5-5"><div class="lab-step">Once the two-minute time limit is up, all matings must stop. 
</div></li><li data-sub-note-item="5-6"><div class="lab-step">The instructor should tally all mating events.
</div></li><li data-sub-note-item="5-7"><div class="lab-step">Find and return the gametes to your ID holder. Give your ID holder to the instructor and notify them if any gametes or supplies are missing. 
</div></li></ul></li><li data-note-item="6" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="6-1"><div class="lab-step">To analyze the data generated during the mating activities, first split up into groups of four. 
</div></li><li data-sub-note-item="6-2"><div class="lab-step">Collect the class data from the instructor for ONE of the four rounds of mating. Note: In this activity, there were several types of females and males distinguished by different letters. These letter types correspond to specific combinations of quality and attractiveness scores. In the compiled data, the average number of matings and reproductive success has already been sorted by letter type for you by the instructor. 
</div></li><li data-sub-note-item="6-3"><div class="lab-step">Use these data to generate four graphs for the round you were assigned. First plot two graphs for the average number of matings against female and male type. 
</div></li><li data-sub-note-item="6-4"><div class="lab-step">Then plot the reproductive success for males and females against male or female type, giving a total of four graphs.
</div></li><li data-sub-note-item="6-5"><div class="lab-step">Once these graphs are complete, each group should present their data to the class, making sure to state the original hypothesis for your assigned round and whether or not the hypothesis was upheld.
</div></li><li data-sub-note-item="6-6"><div class="lab-step">After each group presents, discuss the overall results of the four rounds together. 
</div></li><li data-sub-note-item="6-7"><div class="lab-step">Note the affect that hiding the quality score had on the overall quality of mates chosen.
</div></li><li data-sub-note-item="6-8"><div class="lab-step">Note how hiding or revealing the attractiveness score affected the results.
</div></li><li data-sub-note-item="6-9"><div class="lab-step">Discuss the effect of male parental investment on the number and overall quality of mating pairs.
</div></li></ul></li></ol>

性選択と配偶者選択

Sexual Selection Class Simulation
ExpandNote: In this activity, you and your classmates will portray a sexually reproducing population consisting of an ...

Animal Behavior

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Making Choice Chambers
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">Before beginning the experiments, use one of the Sharpies from the lab table to trace a line around the circumference of the bottom of both bottles. One line should be 1 cm from the base of the bottle and the other should be 1.5 cm from the base of the bottle.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Use a razor blade to make a small cut in each bottle at the mark. 
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Then, insert the scissors and carefully cut along the line until the bottoms of the bottles are removed completely.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Fit one bottle bottom into the other.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Tape the bottles together to make a secure chamber.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">Use the razor blade and scissors to poke a hole into the center of the chamber. 
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Take a piece of tape and fold the edge onto itself to make a tab and place the tape over the hole.  Note: The tab helps with easy removal and reattachment of the tape to the chamber.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">Use the Sharpie to label one side of the chamber A and the other side B.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">Upon receiving the colony of <em>Drosophila</em>, perform a count to make sure there are 40 living individuals in the vial.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Geotaxis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">To perform a geotaxis assay, first attach a clean, empty bottle cap to each end of the choice chamber and turn the fly vial upside down onto its lid.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Open the tape flap in the center of the choice chamber.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Insert the funnel into the hole and tap all of the flies into the chamber. HYPOTHESES: The alternate hypothesis is that more flies will be moving against gravity, or up, regardless of the orientation of the choice chamber. Alternatively, if the flies are positively geotactic, then more flies will move down, or with gravity. The null hypothesis is that the flies will have no preference with regard to gravity and will move randomly within the chamber. 
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Once the <em>Drosophila</em> are in the chamber, close the tape flap, rotate the chamber so that it stands vertically with side A on the bottom, and start the timer.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Immediately, count the number of <em>Drosophila</em> that are moving up and the number that are moving down. Record these data for time point 0 in the table. 
</div></li><li data-sub-note-item="2-6"><div class="lab-step">After 30 seconds, rotate the chamber so that side B is on the bottom and count the number of <em>Drosophila</em> that are moving up or down. Record these data for time point 30 in the table.
</div></li><li data-sub-note-item="2-7"><div class="lab-step">After 60 seconds, rotate the chamber so that side A is on the bottom again and record the number of moving flies for time point 60.  <a href="https://www.jove.com/CDNSource/lm/labs/15/Table_1_Animal_behavior.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Hold the chamber over the fly vial and carefully open the cap to return all of the <em>Drosophila</em> back to their empty vial. 
</div></li></ul></li><li data-note-item="3" data-parent-collapse="">Phototaxis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="3-1"><div class="lab-step">To perform a phototaxis analysis, first cover side A of the choice chamber with tin foil, taking care that the foil reaches to the exact center between the two sides. 
</div></li><li data-sub-note-item="3-2"><div class="lab-step">Ensure the caps are secured on both ends of the chamber.  HYPOTHESES: In this exercise, the alternative hypothesis is that more flies will be found in the light side of the choice chamber, if the flues are attracted to light, or on the dark side if they are repelled by light. The null hypothesis is that <em>Drosophila</em> will have no preference with regard to either stimulus. Therefore, equal numbers of flies will be found on either side of the choice chamber.
</div></li><li data-sub-note-item="3-3"><div class="lab-step">Open the tape flap in the center of the choice chamber, insert the funnel into the hole, and tap all 40 flies into the chamber as previously described.
</div></li><li data-sub-note-item="3-4"><div class="lab-step">Set the timer to sound an alarm at 10 minutes.
</div></li><li data-sub-note-item="3-5"><div class="lab-step">When the alarm sounds, stop the timer, and count the number of flies in the open side B of the chamber.
</div></li><li data-sub-note-item="3-6"><div class="lab-step">Subtract that number from the total number of flies added to the choice chamber to calculate the number of flies in the covered side A.
</div></li><li data-sub-note-item="3-7"><div class="lab-step">Record these data in the table for time point 10.  <a href="https://www.jove.com/CDNSource/lm/labs/15/Table_2_Animal_behavior.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 2</a>
</div></li><li data-sub-note-item="3-8"><div class="lab-step">Carefully, return all of the flies back to their empty vial.
</div></li></ul></li><li data-note-item="4" data-parent-collapse="">Chemotaxis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="4-1"><div class="lab-step">HYPOTHESES: In this exercise, the alternative hypothesis is that more flies will be found on the side of the chamber with the apple cider vinegar if the flies are attracted to the odor. Or, if the flies are repelled by the vinegar, then more of them will be found on the side of the chamber presenting the odorless water. The null hypothesis is that <em>Drosophila</em> will have no preference with regard to either stimulus and will move randomly in the chamber.  Use an individual disposable plastic pipette to dispense two drops of the apple cider vinegar onto one cotton ball for side A of the chamber. 
</div></li><li data-sub-note-item="4-2"><div class="lab-step">Use an individual disposable plastic pipette to dispense two drops of the distilled water onto a cotton ball for side B of the chamber. 
</div></li><li data-sub-note-item="4-3"><div class="lab-step">Fold two pieces of tape over themselves to create individual pieces of double-sided tape and use the tape to attach each cotton ball to the inside of the single bottle cap.
</div></li><li data-sub-note-item="4-4"><div class="lab-step">Screw the bottle caps with the cotton balls onto their corresponding sides of the chamber.
</div></li><li data-sub-note-item="4-5"><div class="lab-step">Transfer all of the flies into the chamber and set the timer to sound an alarm after 10 minutes. 
</div></li><li data-sub-note-item="4-6"><div class="lab-step">When the alarm sounds, stop the timer and record the number of flies in each side of the chamber. <a href="https://www.jove.com/CDNSource/lm/labs/15/Table_3_Animal_behavior.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 3</a>
</div></li><li data-sub-note-item="4-7"><div class="lab-step">At the end of the final experiment, funnel the flies into a morgue tube containing 70% ethanol. 
</div></li><li data-sub-note-item="4-8"><div class="lab-step">Dismantle the choice chambers for recycling.  NOTE: Since all of the chemicals used in these experiments are lab safe, the liquids can be disposed of down the sink, the chambers rinsed, and the solids can be thrown away in the trash. 
</div></li></ul></li><li data-note-item="5" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="5-1"><div class="lab-step">Gather all of the data collected for the geotaxis, phototaxis, and chemotaxis experiments.
</div></li><li data-sub-note-item="5-2"><div class="lab-step">Plot these values as bar graphs for different conditions and different time points.
</div></li><li data-sub-note-item="5-3"><div class="lab-step">Use these data to draw inferences about the behavior of <em>drosophila melanogaster</em>. 
</div></li></ul></li></ol>

Animal Behavior: Studying Taxis Behavior of Drosophila - Procedure

動物の行動

Making Choice Chambers
ExpandBefore beginning the experiments, use one of the Sharpies from the lab table to trace a line around the circumference of the ...

Evolutionary Relationships

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Forming an Evolutionary Hypothesis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">NOTE: A cladogram is an important tool for forming an evolutionary hypothesis. A cladogram is a tree-shaped chart used to depict the hypothetical genealogical relationships between species. The tips or leaves of the chart represent specific species and the branches of the tree are different lengths. The different lengths represent the degree of change between each of the species. The common ancestor of all of the species that a specific line branches from is located at a node, where the branches intersect. Species that share a node are called a sister group.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">To begin, look at the provided blank cladogram and make a hypothesis about where the assigned animals should be placed on the tree. If a blank cladogram has not been provided please <a target="_blank" href="/files/ftp_upload/10562/Blank_Cladogram.png">download one here</a>.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Next, observe the image of the fossilized animal.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Using the morphology, or physical characteristics, of the fossil, make a hypothesis about where on your cladogram it should be placed. Add a new line or mark to indicate your proposed position for the fossil.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Comparing Gene Sequences Using BLAST
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">NOTE: Historically, cladograms have been constructed through the comparison of morphology. Now, genetic sequences can also be compared between species of interest to construct cladograms. DNA sequences are not usually preserved in fossils, so for this lab you will use the BLAST database to compare the DNA sequences of living species that are closely related to the fossil to thousands of other living species.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Before beginning this lab, your instructor will have downloaded the three preserved gene sequences to a folder on your desktop.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Navigate to the BLAST homepage, which the instructor should have left open in your browser, and click on Saved Strategies from the menu at the top of the page.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Click Open on the Evolutionary_Relationships_Gene1.txt file.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Then, click View to go to the Standard Nucleotide BLAST search page. You should be able to see the DNA sequence in the Query Sequence box at the top-right of the screen.
</div></li><li data-sub-note-item="2-6"><div class="lab-step">Scroll to the bottom of the screen and click the BLAST button. Note: It will take a few moments to process and analyze the data. 
</div></li><li data-sub-note-item="2-7"><div class="lab-step">When the data has been processed, a graphic summary will appear.  NOTE: Your query sequence is represented by the blue line at the top of the box. Each of the lines below the blue line represents another sequence in the BLAST database that matches your query. The length and location of these bars represent where and how much of the sequence match up to the query. The color of the bar represents its score, or how identical the sequence is to the query. Below the graphic summary is the Sequences producing significant alignments list, which contains descriptions of the DNA sequences retrieved from the database that most closely align with the query sentence. These are listed in descending order, from the most similar at the top to the least similar at the bottom. On the right, there are several statistics on how closely related each database sequence is to the experimental sequence. The higher the Max, Total, Query Coverage, and Identity scores are, the more similar the query and retrieved sequences are to each other. Similarly, the lower the E value number, the less likely the sequences match was found by random chance and more significantly the alignment is accurate. An E value of 0 indicates a highly significant match.
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Click on the name of the description of the most similar alignment listed. This will take you further down the page to the exact DNA sequence alignment between the retrieved sequence and the query sequence.
</div></li><li data-sub-note-item="2-9"><div class="lab-step">Click on the Sequence ID number. This will open a new tab with more specific information on the retrieved gene sequence.
</div></li><li data-sub-note-item="2-10"><div class="lab-step">Identify and record the scientific and common names of the organism, which should be listed next to Source.
</div></li><li data-sub-note-item="2-11"><div class="lab-step">Identify and record the protein that the gene codes for, which should be listed next to Definition.
</div></li><li data-sub-note-item="2-12"><div class="lab-step">Close the tab and hit Back to return to the Sequences producing significant alignments list.
</div></li><li data-sub-note-item="2-13"><div class="lab-step">Then, repeat steps 7 &#8211; 9 for the next most similar sequences.
</div></li><li data-sub-note-item="2-14"><div class="lab-step">After collecting several species names, return to the Sequences producing significant alignments list and click Select: All to check all of the boxes next to the names of each listed alignment sequence.
</div></li><li data-sub-note-item="2-15"><div class="lab-step">Click on Distance of tree results to create a phylogram based on the similarity of all of the BLAST result gene sequences to the query gene sequence.  Note: Your query fossil relative sequence will be highlighted in yellow.
</div></li><li data-sub-note-item="2-16"><div class="lab-step">Record your sequence&#8217;s location in relation to the other taxon groups shown in the tree.
</div></li><li data-sub-note-item="2-17"><div class="lab-step">Finally, return to the Saved Strategies tab and upload the next saved gene sequence.
</div></li><li data-sub-note-item="2-18"><div class="lab-step">Query the remaining two fossil relative DNA sequences as previously demonstrated (steps 1 &#8211; 16).
</div></li></ul></li><li data-note-item="3" data-parent-collapse="">Data Analysis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="3-1"><div class="lab-step">Based on your gene sequence results, come up with a second hypothesis about where your fossil specimen fit in the original cladogram.
</div></li><li data-sub-note-item="3-2"><div class="lab-step">Compare this placement with your first hypothesis.
</div></li><li data-sub-note-item="3-3"><div class="lab-step">Compare your tree with your classmates&#8217;. If any of your classmates put the fossil on a different branch, ask them how they came to that decision.
</div></li></ul></li></ol>

Evolutionary Relationships: Using BLAST to Test Evolutionary Hypotheses - Procedure

進化的関係

Forming an Evolutionary Hypothesis
ExpandNOTE: A cladogram is an important tool for forming an evolutionary hypothesis. A cladogram is a tree-shaped chart ...

進化論

Macromolecules

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Identifying Macromolecules with Indicator Solutions
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">HYPOTHESES: In this activity, the experimental hypothesis might be that the addition of an indicator dye to each test solution will result in a color change if the test solution contains the macromolecule of interest. The null hypothesis might be that the addition of an indicator dye will not result in a color change in any of the test solutions.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Place 16 clean test tubes in a tube rack in four rows of four tubes and label each row A through D and each column 1 - 4.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Label each test tube with its corresponding position in the test tube rack.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">After this, stir solution A thoroughly and use a pipette to add 4 mL of solution A to each tube in Row A.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Next, stir solution B until it is well mixed and use a new pipette to add 4 mL of solution B to each tube in Row B.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">After stirring solution C thoroughly, add 4 mL of solution C to each tube in Row C.
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Then add 4 mL of soybean-derived vegetable oil to each tube in Row D.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">When all of the tubes have been filled with their respective test solutions, add 200 mL of water to a 400 mL beaker on a hotplate and set the hotplate to high.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">To perform the Benedict&#39;s Test, add 2 mL of Benedict&#39;s reagent to test tubes A1, B1, C1, and D1 while the water is warming.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">Observe the initial color of the tubes.
</div></li><li data-sub-note-item="1-11"><div class="lab-step">When the water begins to boil, transfer each test tube to the beaker and heat the tubes in the boiling water bath for 3 - 5 minutes.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">Then, record the final color of each of the tubes in the appropriate cells of the table and turn off the hotplate. <a href="https://www.jove.com/CDNSource/lm/labs/22/Table_1_Macromolecules.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="1-13"><div class="lab-step">To perform an Iodine-Potassium-Iodide Test, first observe the color of the solutions in the test tubes in column 2.
</div></li><li data-sub-note-item="1-14"><div class="lab-step">Then, add 1 mL of iodine potassium iodide reagent to tubes A2 to D2 and record the final color change in each of the tubes.
</div></li><li data-sub-note-item="1-15"><div class="lab-step">For the Sudan IV Test, observe and record the color of the solutions in the test tubes in column 3.
</div></li><li data-sub-note-item="1-16"><div class="lab-step">Then, add 2 - 3 drops of Sudan IV reagent to test tubes A3 through D3 and record the color change, if any, in each tube.
</div></li><li data-sub-note-item="1-17"><div class="lab-step">For the Biuret Test, observe the color of the solutions in the test tubes in column 4.
</div></li><li data-sub-note-item="1-18"><div class="lab-step">Then, add 1 mL of Biuret reagent to test tubes A4 through D4 and record the final color in each of the tubes.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">Using the results from the indicator tests, each of the four original solutions can be identified as a sugar, starch, lipid, or protein. Refer to the Concepts video to help you determine which tubes contain which macromolecules.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">First, look at the results from the Benedict&#39;s Test. Determine which of the test solutions contain sugar based on your results.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Next, create a bar graph with your five plant species listed on the x-axis and the surface area in cm indicated on the y-axis.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Next, look at the results from the Iodine-Potassium-Iodide Test. Determine which of the test solutions contain starch.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Now, review the results from the Sudan IV Test. Using these observations, determine if any of the test solutions contain lipids.
</div></li><li data-sub-note-item="2-6"><div class="lab-step">Finally, look at the results from the Biuret Test. Determine if any of the test solutions contain proteins.
</div></li></ul></li></ol>

Macromolecules: Identification Using Indicator Solutions - Procedure

高分子

Identifying Macromolecules with Indicator Solutions
ExpandHYPOTHESES: In this activity, the experimental hypothesis might be that the addition of an ...

基礎

Physiology of the Circulatory System

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Measuring Circulatory System Function in Humans
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">Before starting the experiment go to the front of the room and collect an alcohol swab, a sphygmomanometer, and a stethoscope.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Clean the earpieces with the alcohol swab and then insert them into your ears.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">To check if the stethoscope is in the on position, gently tap the flat metal disc called the diaphragm.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">If you hear a sound it&#39;s on. If not, turn the metal disc and tap again and keep turning and tapping the disc until you hear a sound. NOTE: For this experiment you will work in pairs. One person will measure their partner&#39;s blood pressure, while the person being measured will count their heartbeats over the course of one minute.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Fill out table one with the name of the person being monitored. HYPOTHESES: The experimental hypothesis is that systolic blood pressure will be higher when the person is sitting when compared to lying down because more pressure is required for blood to be moved around the body. The null hypothesis is that there will be no difference in systolic blood pressure whether a person is sitting or lying down.<a href="https://www.jove.com/CDNSource/lm/labs/9/Table_1_Physiology_of_Circulatory_System.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="1-6"><div class="lab-step">The person being monitored should begin by lying down. Next, they will need to locate their pulse by placing two fingers next to their windpipe. If a throbbing sensation is felt, this is the pulse.
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Now, ready a time keeping device to count out 1 minute as the person taking their pulse will be having their blood pressure taken at the same time.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">At this point, the person measuring blood pressure should place the cuff of the sphygmomanometer around the other arm of the subject and the stethoscope earpieces into their own ears.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">Start the timer.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">The person being monitored should begin counting the number of heartbeats felt over one full minute. While the subject is taking their pulse, the person taking blood pressure should place the diaphragm of the stethoscope under the cuff on the inside of their partner&#39;s elbow.
</div></li><li data-sub-note-item="1-11"><div class="lab-step">Then, pump the cuff to 200 mm of mercury.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">Gently and slowly twist the metal knob on the sphygmomanometer to release air from the cuff.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">As pressure is released the first Korotkoff sound should be heard. The corresponding value on the sphygmomanometer is the systolic blood pressure. Consistent sounds should be heard until they die out with the last sound signifying the diastolic blood pressure.
</div></li><li data-sub-note-item="1-14"><div class="lab-step">Record the systolic pressure, the diastolic pressure, and the heart rate in Table 1.
</div></li><li data-sub-note-item="1-15"><div class="lab-step">Now, repeat the blood pressure and pulse measurements with each partner keeping the same role but this time the subject should be sitting upright.
</div></li><li data-sub-note-item="1-16"><div class="lab-step">Record these data in the appropriate places in Table 1.
</div></li><li data-sub-note-item="1-17"><div class="lab-step">Finally, the partners should switch spots. So now, the person measuring blood pressure should be taking their pulse and acting as the subject.  NOTE: Be sure to wipe the earpieces of the stethoscope between users and don&#39;t forget to record your measurements in Table 1.
</div></li><li data-sub-note-item="1-18"><div class="lab-step">After finishing the experiment, discard all alcohol swabs into the trash. Then, bring the sphygmomanometer and stethoscope back to the front of the room.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Measuring Respiratory Rate in Fish
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">To prepare for this second experiment, collect two clear containers.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Begin by filling half of one container with aquarium water.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Then, use a fish net to gently catch a goldfish and place it into the cup. Pinch the net around the fish or else it may jump out of the net.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Familiarize yourself with the operculum of the fish. The operculum covers the gills and it is located behind and below the eye. When the fish respires the operculum moves in and out and this is one gill beat. HYPOTHESES: In this experiment you will measure the gill beats of the fish in different water temperatures. The hypothesis is that there will be a significant correlation between the temperature and respiration rate in goldfish. The null hypothesis is that there will be no correlation between temperature and respiration rate.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Take the other empty container with a screw cap, the cup with your fish, and the net to the water tanks.
</div></li><li data-sub-note-item="2-6"><div class="lab-step">Then, fill the empty container with the water from the lowest temperature water bath. This will act as your first respiration rate chamber.
</div></li><li data-sub-note-item="2-7"><div class="lab-step">Next, hold the fish net over a wastewater container and dump out the water from the cup holding the fish so that the fish falls into the net. Pinch the net around the fish without touching the fish itself.
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Bring the net over the top of the container with the bath water and let go of the net to release the fish into the vial.
</div></li><li data-sub-note-item="2-9"><div class="lab-step">Once the fish is in the container, submerge it back into the water bath and then screw the cap on to prevent outside oxygen from entering the vial.
</div></li><li data-sub-note-item="2-10"><div class="lab-step">Next, remove the container from the controlled temperature water bath and place it on your workbench surface to avoid warming the water with body heat.
</div></li><li data-sub-note-item="2-11"><div class="lab-step">One partner should immediately begin counting the gill beats as soon as the second partner starts the stopwatch.
</div></li><li data-sub-note-item="2-12"><div class="lab-step">Continue counting the beats for 20 seconds. Record the number of beats in column A of Table 2. <a href="https://www.jove.com/CDNSource/lm/labs/9/Table_2_Physiology_of_Circulatory_System.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 2</a>
</div></li><li data-sub-note-item="2-13"><div class="lab-step">Repeat the measurement one more time and record the number in column B.
</div></li><li data-sub-note-item="2-14"><div class="lab-step">Next, hold the fish net over a wastewater container and dump out the water from the fish vial so that the fish falls into the net.  IMPORTANT: Move from lowest to highest temperature, so the fish is not stressed between trials.
</div></li><li data-sub-note-item="2-15"><div class="lab-step">Repeat the experiment for the remaining four temperature conditions (steps 8 &#8211; 13).
</div></li><li data-sub-note-item="2-16"><div class="lab-step">Record all the values for different temperatures in Table 2.
</div></li><li data-sub-note-item="2-17"><div class="lab-step">Once you are done, slowly mix water from the last respiration chamber with the fish tank water and then return the fish to the home tank. NOTE: This should be done over the course of approximately 10 minutes.
</div></li></ul></li><li data-note-item="3" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="3-1"><div class="lab-step">It&#39;s time to analyze the data that you just collected. Look at the blood pressure and heart rate measurements for the different trials. If they were different, make a hypothesis to explain your observations. 
</div></li><li data-sub-note-item="3-2"><div class="lab-step">For the gill beat experiment, average the numbers in columns A and B and calculate the fish respiration rate as gill beats per minute.
</div></li><li data-sub-note-item="3-3"><div class="lab-step">Next, plot the graph of temperature versus gill beats per minute.
</div></li><li data-sub-note-item="3-4"><div class="lab-step">Record any correlations you see between the variables. If a correlation is observed, determine if it is positive or negative and provide an explanation for this result.  NOTE: Consider the fact that fishes are ectotherms and the impact of temperature on dissolved oxygen.
</div></li></ul></li></ol>

循環器系の生理学

Measuring Circulatory System Function in Humans
ExpandBefore starting the experiment go to the front of the room and collect an alcohol swab, a ...

Artificial Selection

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Observing Artificial Selection in Plants 
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">To begin the experiment, collect five two-week old grown plants from the generation one population that appear healthy. Note: Plants should appear green, not wilted, and not have damaged leaves. Experimental Hypothesis: The experimental hypothesis for this lab might be that the number of lead trichomes varies naturally, but the average number will increase over generations if only plants with the highest number of trichomes are interbred or decrease if those with the least are interbred. Null Hypothesis: The null hypothesis might be that the average number of trichomes will not change over subsequent generations after artificial selection.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Label each plant with a letter from A to E and with both partners&#8217; names.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Record any natural differences you see between your plant samples. Note: All of the plants are the same age, so any observed variation is due to expressed phenotype, not development.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Use a hand lens to count the number of trichomes found on the petiole of the lowest leaf, not including the round, first-grown cotyledons for each plant. Note: It can be helpful to shine a light onto the plant and to hold it against a dark background, such as a lab bench, during the trichome analysis. 
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Both partners will conduct counts on each plant independently and compare answers. If the counts are different, enlist a third counter or take the average of the two results.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">After this, determine the median number of trichomes for the entire class.  <a target="__blank" style="display:block;padding:20px;" href="https://www.jove.com/CDNSource/lm/labs/2/Lab_2_Table_A.xlsx">Click Here to download Table A</a>
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Determine the top and bottom 10% of the plants from the class trichome count. These groups will make up the two artificially selected lines: high hairiness and low hairiness, respectively.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">If more than 10% of the plants have no trichomes, select 10% of those plants randomly for the low hairiness plant line.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">Separate the two groups of selected plants and label them clearly.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">Move the plants to a well-lit area and keep them well watered until they develop flowers. Note: Flowers typically develop around 14 days.
</div></li><li data-sub-note-item="1-11"><div class="lab-step">When the flowers are fully developed, use a pollination wand or paintbrush to rub the stigma and anther of one plant&#8217;s flowers for a few seconds.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">Using the same wand or brush, rub the inside of each flower of every other plant in the same selection line.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">Using a different wand or brush, repeat steps 11 through 12 on the other line. Note: Take care not to cross the lines. Spread the pollen only among the appropriately selected plants.
</div></li><li data-sub-note-item="1-14"><div class="lab-step">Return the generation one plants to the growing areas. When the plants reach approximately 1 month old, they should produce mature seed pods.
</div></li><li data-sub-note-item="1-15"><div class="lab-step">Allow the plants to dry out once the seed pods turn yellow.
</div></li><li data-sub-note-item="1-16"><div class="lab-step">Break the seed pods open and collect the seeds.
</div></li><li data-sub-note-item="1-17"><div class="lab-step">Obtain new pots for the generation two seeds.
</div></li><li data-sub-note-item="1-18"><div class="lab-step">Fill the pots with moist potting soil about halfway and add a fertilizer pellet to each pot.
</div></li><li data-sub-note-item="1-19"><div class="lab-step">Fill the pots with more soil to fill just below the lip.
</div></li><li data-sub-note-item="1-20"><div class="lab-step">Make a shallow depression in the soil of each pot with a probe.
</div></li><li data-sub-note-item="1-21"><div class="lab-step">Touch a clean toothpick to a gluestick and use the toothpick to transplant the seeds into the soil one by one.
</div></li><li data-sub-note-item="1-22"><div class="lab-step">Cover the seeds with a thin layer of soil.
</div></li><li data-sub-note-item="1-23"><div class="lab-step">Label the pots with their hairiness line and place them in the designated growing area.
</div></li><li data-sub-note-item="1-24"><div class="lab-step">Use a plastic dropper to water the plants as needed to ensure they do not dry out.
</div></li><li data-sub-note-item="1-25"><div class="lab-step">When the plants are two weeks old, collect five plants from each line and label them A to E.
</div></li><li data-sub-note-item="1-26"><div class="lab-step">Then, count the trichomes on these plants as previously described (steps 4- 5).
<a target="__blank" style="display:block;padding:20px;" href="https://www.jove.com/CDNSource/lm/labs/2/Lab_2_Table_B.xlsx">Click Here to download Table B</a>
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Data Analysis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">Create a frequency graph based on the generation one class data with the trichome count on the x axis and the number of plants on the y axis. Label the mean. 
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Then, create frequency graphs for both of the selected hairiness lines.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Compare the frequency graphs for generation one and both hairiness lines.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Take note of phenotypic shifts observed in the selected hairiness lines.
</div></li></ul></li></ol>

Artificial Selection: Visualization in Plants and Data Analysis - Procedure

人工選択

Observing Artificial Selection in Plants 
ExpandTo begin the experiment, collect five two-week old grown plants from the generation one population that ...