この作品は、NIH グラント GM59354 シャロン E. Bickel に授与によって支えられました。Q. ウイ グエンにアン Lavanway 共焦点顕微鏡のヘルプとテクニカル サポートに J. エミリアーノ リード蛍光腕プローブの生成のためのプロトコルの開発に支援ありがちましょう。また、有用な議論やアドバイスのためのショウジョウバエのコミュニティで多くの同僚らを感謝いたします。

<ol>
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L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Defective+cohesin+is+associated+with+age-dependent+misaligned+chromosomes+in+oocytes.">Defective cohesin is associated with age-dependent misaligned chromosomes in oocytes.</a> Reprod Biomed Online. 16 (1), 103-112 (2008).</li><li>Subramanian, V. V., Bickel, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Aging+predisposes+oocytes+to+meiotic+nondisjunction+when+the+cohesin+subunit+SMC1+is+reduced.">Aging predisposes oocytes to meiotic nondisjunction when the cohesin subunit SMC1 is reduced.</a> PLoS Genet. 4 (11), e1000263 (2008).</li><li>Chiang, T., Duncan, F. E., Schindler, K., Schultz, R. M., Lampson, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evidence+that+weakened+centromere+cohesion+is+a+leading+cause+of+age-related+aneuploidy+in+oocytes.">Evidence that weakened centromere cohesion is a leading cause of age-related aneuploidy in oocytes.</a> Curr. Biol. 20 (17), 1522-1528 (2010).</li><li>Lister, L. 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=Age-related+meiotic+segregation+errors+in+mammalian+oocytes+are+preceded+by+depletion+of+cohesin+and+Sgo2.">Age-related meiotic segregation errors in mammalian oocytes are preceded by depletion of cohesin and Sgo2.</a> Curr. Biol. 20 (17), 1511-1521 (2010).</li><li>Duncan, F. E., 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=Chromosome+cohesion+decreases+in+human+eggs+with+advanced+maternal+age.">Chromosome cohesion decreases in human eggs with advanced maternal age.</a> Aging Cell. 11 (6), 1121-1124 (2012).</li><li>Yun, Y., Lane, S. I., Jones, K. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Premature+dyad+separation+in+meiosis+II+is+the+major+segregation+error+with+maternal+age+in+mouse+oocytes.">Premature dyad separation in meiosis II is the major segregation error with maternal age in mouse oocytes.</a> Development. 141 (1), 199-208 (2014).</li><li>Subramanian, V. V., Bickel, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heterochromatin-mediated+association+of+achiasmate+homologues+declines+with+age+when+cohesion+is+compromised.">Heterochromatin-mediated association of achiasmate homologues declines with age when cohesion is compromised.</a> Genetics. 181, 1207-1218 (2009).</li><li>Jeffreys, C. A., Burrage, P. S., Bickel, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+model+system+for+increased+meiotic+nondisjunction+in+older+oocytes.">A model system for increased meiotic nondisjunction in older oocytes.</a> Curr. Biol. 13 (6), 498-503 (2003).</li><li>Weng, K. A., Jeffreys, C. A., Bickel, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rejuvenation+of+Meiotic+Cohesion+in+Oocytes+during+Prophase+I+Is+Required+for+Chiasma+Maintenance+and+Accurate+Chromosome+Segregation.">Rejuvenation of Meiotic Cohesion in Oocytes during Prophase I Is Required for Chiasma Maintenance and Accurate Chromosome Segregation.</a> PLoS Genetics. 10 (9), e1004607 (2014).</li><li>Perkins, A. T., Das, T. M., Panzera, L. C., Bickel, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oxidative+stress+in+oocytes+during+midprophase+induces+premature+loss+of+cohesion+and+chromosome+segregation+errors.">Oxidative stress in oocytes during midprophase induces premature loss of cohesion and chromosome segregation errors.</a> Proc Natl Acad Sci U S A. 113 (44), E6823-E6830 (2016).</li><li>Dernburg, A. F., Sedat, J. W., Hawley, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Direct+evidence+of+a+role+for+heterochromatin+in+meiotic+chromosome+segregation.">Direct evidence of a role for heterochromatin in meiotic chromosome segregation.</a> Cell. 86, 135-146 (1996).</li><li>Dernburg, A. F., Sedat, J. 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著者は競合する金銭的な利益を宣言しません。

私と中期 prometaphase の腕の結束の状態を評価するプローブの魚の使用私のショウジョウバエの卵母細胞はショウジョウバエの減数分裂の分野の重要な進歩。歴史的に、ショウジョウバエ研究者は成熟卵11,18,19腕結束の早期喪失を推論するための遺伝子検査に限られています。今、紹介した方法で腕の結?...

有糸分裂と減数分裂時の染色体の分離が適切な必要とその姉妹染色分け体の凝集性の確立、維持、・連携1,2のリリースします。凝集度は S 期の間に確立され、姉妹を保持する物理の連携を形成するコヒーシン複雑、により媒介される染色分け体一緒に。減数分裂、クロス オーバーの遠位結束も組換え同族体を一緒に保持する機能し、この物理的な関連付けにより、適切な方向 (図 1)3,4、減数分裂に 2価の紡錘 5。リリース アームの後期で結束対極スピンドルに分離する同族体が可能します。ただし、腕結束がある場合途中で失われた組換え同族体の物理的な接続を失う、異数性配偶子 (図 1) につながることができます、ランダムに分離します。
人間の卵子の染色体分配のエラー6、ダウン症候群などの先天異常と流産の原因、その発生率は、母体年齢7とともに指数関数的に増加します。胎児卵母細胞で姉妹染色分け体の凝集が確立され、出生前に減数分裂組み換えが完了しました。卵母細胞、中期前期私排卵までと逮捕この逮捕中に、組換え同族体の継続的な物理的な連合の依存姉妹染色分け体結束。したがって、減数分裂と正常妊娠の転帰の間に正確な分離は、最大 5 年間の凝集力がそのまま残ること必要があります。
人間の卵子の減数分裂長期逮捕時に結束の早期喪失は母親年齢の効果に貢献する提案されているし、証拠の複数のラインは、この仮説8,9をサポートします。ただし、ヒトの卵母細胞の減数分裂の結束を研究の課題を考えると、この現象の私達の理解の多くは、モデル生物5,10、11,12,の使うことで13,14,15。
ショウジョウバエの卵母細胞は減数分裂の結束と染色体の偏析の研究のための多数の利点を提供しています。単純な遺伝的アッセイ異数性配偶子から子孫を回復し、 Xの忠実度を測定することができます-大規模な11,16,17染色体分離。さらに、1 つも減数分裂期組換え同族体 missegregate アーム結束11,18,の早期喪失と一貫性のある表現型、私ために染色体分離エラーが発生するかどうかを決定可能性があります19. 直接ショウジョウバエの卵母細胞の減数分裂の結束の状態の観察が可能な蛍光現場の交配 (魚) を使用しても。反復的な衛星シーケンスに交配する蛍光のオリゴヌクレオチドが使用されているため 10 年以上 pericentromeric 結束の成熟したショウジョウバエ卵4、20アームの分析を監視するには凝集度は、はるかに困難されています。腕の結束の状態の可視化には、シングル コピーの大規模な地域にわたるプローブ シーケンスし、アーム結合が存在しないときに個々 の妹のため目に見えるシグナル染色分け体が発生するには十分に明るいですが必要です。さらに、卵母細胞固定条件と分類された Dna のサイズは大きな成熟したショウジョウバエ卵母細胞 (200 μ m 幅 500 μ m 長で) に浸透21を促進する必要があります。最近、腕プローブが正常に利用いる prometaphase で信号の検出できませんでした述べたけど、著者、後期中ショウジョウバエ卵母細胞染色分け体を可視化するまたは中期卵22を逮捕しました。ここでは、 Xの生成のための詳しいプロトコルを提供-私と中期染色分け体結束 prometaphase の妹の早期喪失の試金する私たちを許可している卵母細胞調製条件と魚プローブ染色体腕私卵。減数分裂の結束の維持に必要な遺伝子の製品を識別するために私たちを有効に、これらのテクニックになります他の姉妹のために試金する染色分け体結束異なった遺伝子型の成熟のショウジョウバエ卵の欠陥します。

<table><tbody><tr><td>&lt;strong&gt;Kits&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Midi Prep kit</td><td>Qiagen</td><td>12143</td><td>Prep BAC clone DNA</td></tr><tr><td>GenomePlex Complete Whole Genome Amplification (WGA) Kit</td><td>Sigma</td><td>WGA2</td><td>Amplify BAC clone DNA</td></tr><tr><td>ARES Alexa Fluor 647 DNA labeling kit</td><td>Invitrogen</td><td>A21676</td><td>Label BAC clone DNA</td></tr><tr><td>PCR purification kit</td><td>Qiagen</td><td>28104</td><td>Remove non-conjugated dye following labeling of BAC clone DNA</td></tr><tr><td>&lt;strong&gt;Name&lt;/strong&gt;</td><td>&lt;strong&gt;Company&lt;/strong&gt;</td><td>&lt;strong&gt;Catalog Number&lt;/strong&gt;</td><td>&lt;strong&gt;Comments&lt;/strong&gt;</td></tr><tr><td>&lt;strong&gt;Chemicals &amp;amp; Solutions&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>&lt;strong&gt;Note: &lt;/strong&gt;All solutions are prepared using sterile ultrapure water and should be sterilized either by autoclave or filter sterilization.</td><td></td><td></td><td></td></tr><tr><td>Bovine serum albumin (BSA)</td><td>Fisher Scientific</td><td>BP1600-100</td><td>Prepare 10% stock&lt;br/&gt; Freeze aliquots</td></tr><tr><td>Calcium chloride</td><td>Fisher Scientific</td><td>C75-500</td><td></td></tr><tr><td>DAPI (4&amp;rsquo;, 6-Diamidino-2-Phenylindole, Dihydrochloride)</td><td>Invitrogen</td><td>D1306</td><td>&lt;strong&gt;Toxic:&lt;/strong&gt; wear appropriate protection. Prepare 100&amp;micro;g/ml stock in 100% ethanol, store in aliquots at -20 &amp;deg;C. Prepare 1 &amp;micro;g/ml solution in 2X SSCT before use.</td></tr><tr><td>Dextran sulfate</td><td>Sigma</td><td>D-8906</td><td></td></tr><tr><td>Drierite</td><td>Drierite Company</td><td>23001</td><td></td></tr><tr><td>Dithiothreitol (DTT)</td><td>Invitrogen</td><td>15508-013</td><td>Prepare 10 mM stock</td></tr><tr><td>dTTP (10 &amp;micro;mol, 100 &amp;micro;l)</td><td>Boehringer Mannheim</td><td>1277049</td><td>Prepare 1 mM stock</td></tr><tr><td>EDTA (Disodium ethylenediamine tetraacetic acid)</td><td>Fisher Scientific</td><td>S311-500</td><td>Prepare 250 mM stock</td></tr><tr><td>100% ethanol (molecular grade, 200 proof)</td><td>Decon Laboratories</td><td>2716</td><td></td></tr><tr><td>Tris (Ultra Pure)</td><td>Invitrogen</td><td>15504-020</td><td></td></tr><tr><td>EGTA (Ethylenebis(oxyethylenenitrilo)tetraacetic acid)</td><td>Sigma</td><td>E-3889</td><td></td></tr><tr><td>16% formaldehyde</td><td>Ted Pella, Inc.</td><td>18505</td><td>&lt;strong&gt;Toxic:&lt;/strong&gt; wear appropriate protection</td></tr><tr><td>Formamide</td><td>Invitrogen</td><td>AM9342</td><td>&lt;strong&gt;Toxic: &lt;/strong&gt;wear appropriate protection</td></tr><tr><td>Glucose</td><td>Fisher Scientific</td><td>D16-1</td><td></td></tr><tr><td>Glycogen</td><td>Roche</td><td>901393</td><td></td></tr><tr><td>HEPES</td><td>Boehringer Mannheim</td><td>737-151</td><td></td></tr><tr><td>Heptane</td><td>Fisher Scientific</td><td>H350-4</td><td>&lt;strong&gt;Toxic:&lt;/strong&gt; wear appropriate protection.</td></tr><tr><td>Hydrochloric acid</td><td>Millipore</td><td>HX0603-4</td><td>&lt;strong&gt;Toxic:&lt;/strong&gt; wear appropriate protection.</td></tr><tr><td>Hydroxylamine</td><td>Sigma</td><td>438227</td><td>Prepare 3 M stock</td></tr><tr><td>4.9 M Magnesium chloride</td><td>Sigma</td><td>104-20</td><td></td></tr><tr><td>Na&lt;sub&gt;2&lt;/sub&gt;HPO&lt;sub&gt;4 &lt;/sub&gt;&amp;Yuml; 7H&lt;sub&gt;2&lt;/sub&gt;O</td><td>Fisher Scientific</td><td>S373-500</td><td></td></tr><tr><td>NaH&lt;sub&gt;2&lt;/sub&gt;PO&lt;sub&gt;4 &lt;/sub&gt;&amp;Yuml; 2H&lt;sub&gt;2&lt;/sub&gt;O</td><td>Fisher Scientific</td><td>S369-500</td><td></td></tr><tr><td>Poly-L-lysine (0.1mg/ml)</td><td>Sigma</td><td>P8920-100</td><td></td></tr><tr><td>Potassium acetate</td><td>Fisher Scientific</td><td>BP364-500</td><td></td></tr><tr><td>Sodium acetate</td><td>Fisher Scientific</td><td>S209-500</td><td>Prepare 3M stock</td></tr><tr><td>Sodium cacodylate</td><td>Polysciences, Inc.</td><td>1131</td><td>&lt;strong&gt;Toxic: &lt;/strong&gt;wear appropriate protection. Prepare 400mM stock</td></tr><tr><td>Sodium citrate</td><td>Fisher Scientific</td><td>BP327-1</td><td></td></tr><tr><td>Sodium chloride</td><td>Fisher Scientific</td><td>S271-3</td><td>Sodium chloride</td></tr><tr><td>Sucrose</td><td>Fisher Scientific</td><td>S5-500</td><td></td></tr><tr><td>10% Tween 20</td><td>Thermo Scientific</td><td>28320</td><td>Surfact-Amps</td></tr><tr><td>10% Triton X-100</td><td>Thermo Scientific</td><td>28314</td><td>Surfact-Amps</td></tr><tr><td>&lt;strong&gt;Name&lt;/strong&gt;</td><td>&lt;strong&gt;Company&lt;/strong&gt;</td><td>&lt;strong&gt;Catalog Number&lt;/strong&gt;</td><td>&lt;strong&gt;Comments&lt;/strong&gt;</td></tr><tr><td>&lt;strong&gt;Solutions&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>&lt;strong&gt;Note: &lt;/strong&gt;All solutions are prepared using sterile ultrapure water and should be sterilized either by autoclave or filter sterilization.</td><td></td><td></td><td></td></tr><tr><td>TE buffer</td><td></td><td></td><td>10 mM Tris, 1 mM EDTA, pH = 8.0</td></tr><tr><td>20X SSC (Saline Sodium Citrate)</td><td></td><td></td><td>3 M NaCl, 300 mM sodium citrate</td></tr><tr><td>2X cacodylate fix solution</td><td></td><td></td><td>&lt;strong&gt;Toxic:&lt;/strong&gt; wear appropriate protection. 200 mM sodium cacodylate, 200 mM sucrose, 80 mM sodium acetate, 20 mM EGTA</td></tr><tr><td>1.1X Hybridization buffer</td><td></td><td></td><td>3.3X SSC, 55% formamide, 11% dextran sulfate</td></tr><tr><td>Fix solution</td><td></td><td></td><td>&lt;strong&gt;Toxic: &lt;/strong&gt;wear appropriate protection. 4% formaldehyde, 1X cacodylate fix solution</td></tr><tr><td>PBSBTx</td><td></td><td></td><td>1X PBS, 0.5% BSA, 0.1% Trition X-100</td></tr><tr><td>PBSTx</td><td></td><td></td><td>1X PBS, 1% Trition X-100</td></tr><tr><td>Extraction buffer (PBSTx + Rnase)</td><td></td><td></td><td>1X PBS, 1% Trition X-100, 100 &amp;micro;g/mL RNase</td></tr><tr><td>2X SSCT</td><td></td><td></td><td>2X SSC, 0.1% Tween 20</td></tr><tr><td>2X SSCT + 20% formamide</td><td></td><td></td><td>&lt;strong&gt;Toxic:&lt;/strong&gt; wear appropriate protection. 2X SSC, 0.1% Tween 20, 20% formamide</td></tr><tr><td>2X SSCT + 40% formamide</td><td></td><td></td><td>&lt;strong&gt;Toxic:&lt;/strong&gt; wear appropriate protection. 2X SSC, 0.1% Tween 20, 40% formamide</td></tr><tr><td>2X SSCT + 50% formamide</td><td></td><td></td><td>&lt;strong&gt;Toxic:&lt;/strong&gt; wear appropriate protection. 2X SSC, 0.1% Tween 20, 50% formamide</td></tr><tr><td>&lt;strong&gt;Name&lt;/strong&gt;</td><td>&lt;strong&gt;Company&lt;/strong&gt;</td><td>&lt;strong&gt;Catalog Number&lt;/strong&gt;</td><td>&lt;strong&gt;Comments&lt;/strong&gt;</td></tr><tr><td>&lt;strong&gt;Enzymes&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>AluI</td><td>New England Biolabs</td><td>R0137S</td><td></td></tr><tr><td>HaeIII</td><td>New England Biolabs</td><td>R0108S</td><td></td></tr><tr><td>MseI</td><td>New England Biolabs</td><td>R0525S</td><td></td></tr><tr><td>MspI</td><td>New England Biolabs</td><td>R0106S</td><td></td></tr><tr><td>RsaI</td><td>New England Biolabs</td><td>R0167S</td><td></td></tr><tr><td>BfuCI</td><td>New England Biolabs</td><td>R0636S</td><td></td></tr><tr><td>100X BSA</td><td>New England Biolabs</td><td></td><td>Comes with NEB enzymes</td></tr><tr><td>10X NEB buffer #2</td><td>New England Biolabs</td><td></td><td>Restriction enzyme digestion buffer. Comes with NEB enzymes</td></tr><tr><td>Terminal deoxynucleotidyl transferase (TdT) 400 U/&amp;micro;l</td><td>Roche/Sigma</td><td>3333566001</td><td></td></tr><tr><td>TdT buffer</td><td>Roche/Sigma</td><td></td><td>Comes with TdT enzyme</td></tr><tr><td>Cobalt chloride</td><td>Roche/Sigma</td><td></td><td>&lt;strong&gt;Toxic: &lt;/strong&gt;wear appropriate protection. Comes with TdT enzyme</td></tr><tr><td>RNase A (10 mg/mL)</td><td>Thermo-Scientific</td><td>EN0531</td><td></td></tr><tr><td>&lt;strong&gt;Name&lt;/strong&gt;</td><td>&lt;strong&gt;Company&lt;/strong&gt;</td><td>&lt;strong&gt;Catalog Number&lt;/strong&gt;</td><td>&lt;strong&gt;Comments&lt;/strong&gt;</td></tr><tr><td>&lt;strong&gt;Cytology Tools etc.&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Forceps</td><td>Dumont</td><td></td><td>#5 INOX, Biologie</td></tr><tr><td>9&amp;rdquo; Disposable glass Pasteur pipettes</td><td>Fisher</td><td>13-678-20C</td><td>Autoclave to sterilize</td></tr><tr><td>Shallow glass dissecting dish</td><td></td><td></td><td>Custom made</td></tr><tr><td>Deep well dish (3 wells)</td><td>Pyrex</td><td>7223-34</td><td></td></tr><tr><td>Fisherfinest Premium microscope slides</td><td>Fisher Scientific</td><td>22-038-104</td><td>Used to cover deep well dishes</td></tr><tr><td>Frosted glass slides, 25 x 75 mm</td><td>VWR Scientific</td><td>48312-002</td><td></td></tr><tr><td>Glass slides, 3 x 1 in, 1 mm thick</td><td>Thermo-Scientific</td><td>3051</td><td></td></tr><tr><td>Coverslips, 10 x 10 mm, No. 1.5</td><td>Thermo-Scientific</td><td>3405</td><td></td></tr><tr><td>Tungsten needle</td><td></td><td></td><td>homemade</td></tr><tr><td>Prolong GOLD mounting media</td><td>Molecular Probes</td><td>P36930</td><td></td></tr><tr><td>Compressed-air in can</td><td>Various</td><td></td><td></td></tr><tr><td>&lt;strong&gt;Name&lt;/strong&gt;</td><td>&lt;strong&gt;Company&lt;/strong&gt;</td><td>&lt;strong&gt;Catalog Number&lt;/strong&gt;</td><td>&lt;strong&gt;Comments&lt;/strong&gt;</td></tr><tr><td>&lt;strong&gt;Equipment&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>PCR machine</td><td>Various</td><td></td><td></td></tr><tr><td>Nanodrop 2000, spectrophotometer</td><td>Thermo-Scientific</td><td></td><td>microvolume spectrophotometer</td></tr><tr><td>Vortexer</td><td>Various</td><td></td><td></td></tr><tr><td>Table top microfuge at room temperature</td><td>Various</td><td></td><td></td></tr><tr><td>Table top microfuge at 4 &amp;deg;C</td><td>Various</td><td></td><td></td></tr><tr><td>Heat block</td><td>Various</td><td></td><td></td></tr><tr><td>Hybridization oven or incubator with rotator</td><td>Various</td><td></td><td></td></tr><tr><td>Nutator</td><td>Various</td><td></td><td></td></tr><tr><td>A1RSi laser scanning confoal</td><td>Nikon</td><td></td><td>40X oil Plan Fluor DIC (NA 1.3)</td></tr><tr><td>&lt;strong&gt;Name&lt;/strong&gt;</td><td>&lt;strong&gt;Company&lt;/strong&gt;</td><td>&lt;strong&gt;Catalog Number&lt;/strong&gt;</td><td>&lt;strong&gt;Comments&lt;/strong&gt;</td></tr><tr><td>&lt;strong&gt;Consumables etc.&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>50 mL conical tubes</td><td>Various</td><td></td><td></td></tr><tr><td>15 mL conical tubes</td><td>Various</td><td></td><td></td></tr><tr><td>1.5 mL microfuge tubes</td><td>Various</td><td></td><td></td></tr><tr><td>500 &amp;micro;l microfuge tubes</td><td>Various</td><td></td><td></td></tr><tr><td>200 &amp;micro;l PCR tubes</td><td>Various</td><td></td><td></td></tr><tr><td>Plastic container with tight fitting lid</td><td>Various</td><td></td><td>To hold Drierite</td></tr><tr><td>Kimwipes</td><td>Various</td><td></td><td>disposable wipes</td></tr><tr><td>Parafilm</td><td>Various</td><td></td><td>paraffin film</td></tr><tr><td>&lt;strong&gt;Name&lt;/strong&gt;</td><td>&lt;strong&gt;Company&lt;/strong&gt;</td><td>&lt;strong&gt;Catalog Number&lt;/strong&gt;</td><td>&lt;strong&gt;Comments&lt;/strong&gt;</td></tr><tr><td>&lt;strong&gt;Other&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>HPLC purified 5&#039;-labeled oligonucleotides</td><td>Integrated DNA Technologies</td><td></td><td>Cy3-labeled probes that recognize the 359 bp satellite repeat of the &lt;em&gt;X&lt;/em&gt; chromosome</td></tr><tr><td>Volocity 3D Image Analysis Software</td><td>PerkinElmer</td><td></td><td>Version 6.3</td></tr></tbody></table>

using fluorescence in situ hybridization (fish) to monitor the state of arm cohesion in prometaphase and metaphase i drosophila oocytes

ヒトでは、卵母細胞における染色体分離エラーが流産や先天性欠損症の大半を担当です。さらに、女性の年齢、妊娠 aneuploid 胎児が大幅に増加、このような現象のリスクは母体年齢効果と呼ばれます。減数分裂の分裂時の正確な染色体分離の要件の 1 つは妹のメンテナンス卵母細胞が発生する拡張前期期間中に染色分け体結束。人間のモデル生物細胞学的証拠では、老化の過程減数分裂凝集度を劣化させることを示唆しています。人間の卵子に偏析エラーは減数分裂の間に最も普及している、さらに、私は、腕の結束の早期喪失と一貫性のあります。モデル有機体の使用は、凝集度の年齢依存性損失の根底にあるメカニズムの解明にとって重要です。キイロショウジョウバエでは、卵母細胞の減数分裂の凝集の制御を勉強するためのいくつかの利点を提供しています。しかし、最近まで、遺伝テストのみでした腕結束異なった遺伝子型の異なる実験条件下で卵母細胞の損失の分析に利用できます。ここでは、詳細なプロトコルで、私は、ショウジョウバエの卵母細胞を逮捕された中期の腕 prometaphase の結束の交配 (魚) を直接可視化する蛍光その場で使用するため欠陥を提供。X染色体の遠位腕に交配魚プローブを生成および共焦点 Z スタックを収集、研究者が 3 つの次元での個々 の魚信号数を視覚化して姉妹かどうかを決定する染色分け体腕区切られました。手順では、ショウジョウバエの卵母細胞の何百もの arm 結束欠陥を定量化可能です。そのため、このメソッドは、結束のメンテナンスだけでなく、老化プロセスの間に死につながる要因に寄与するメカニズムを調査するため重要なツールを提供します。

X染色体の細胞学的地域 6E 7B に交配腕プローブで得られた画像を図 5に示します。共同の DAPI、ローカライズ信号でこの検出結果の例は、背景から識別しやすいと正常に異なった遺伝子型19腕結束の欠陥を定量化するために使用されています。結束性欠陥の定量化は制限された prometaphase に私と中期私段階;核膜崩壊前に卵?...

Watch this Scientific Journal Video about Using Fluorescence In Situ Hybridization FISH to Monitor the State of Arm Cohesion in Prometaphase and Metaphase I Drosophila Oocytes at JoVE.com

蛍光 In situ ハイブリダイゼーション FISH を使用した前中期および中期 I ショウジョウバエ卵母細胞のアーム凝集状態のモニタリング |発達 |生物学 |ビデオ

Using Fluorescence In Situ Hybridization FISH to Monitor the State of Arm Cohesion in Prometaphase and Metaphase I Drosophila Oocytes

Prometaphase の腕凝集度と中期の状態を監視する蛍光In Situハイブリダイゼーション (魚) を使用して私はショウジョウバエの卵母細胞

この原稿は、 Xを生成するための詳細な方法を提示-染色体腕プローブおよび実行する蛍光その場で交配 (魚) の妹の状態を調べるに染色分け体結束の prometaphase とを逮捕した中期ショウジョウバエ卵。このプロトコルは、減数分裂の腕の結合はそのままか、異なった遺伝子型の崩壊かどうかを決定するのに適しています。

In humans, chromosome segregation errors in oocytes are responsible for the majority of miscarriages and birth defects. Moreover, as women age, their risk ...

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Using Fluorescence In Situ Hybridization (FISH) to Monitor the State of Arm Cohesion in Prometaphase and Metaphase I Drosophila Oocytes

Meiosis

SCIENTISTS IN ACTION

11.3K ビュー.  Dartmouth College. この原稿は、 Xを生成するための詳細な方法を提示-染色体腕プローブおよび実行する蛍光その場で交配 (魚) の妹の状態を調べるに染色分け体結束の prometaphase とを逮捕した中期ショウジョウバエ卵。このプロトコルは、減数分裂の腕の結合はそのままか、異なった遺伝子型の崩壊かどうかを決定するのに適しています。

ビデオ: Using Fluorescence In Situ Hybridization FISH to Monitor the State of Arm Cohesion in Prometaphase and Metaphase I Drosophila Oocytes

Using Mouse Oocytes to Assess Human Gene Function During Meiosis I

Embryonic aneuploidy is the major genetic cause of infertility in humans. Most of these events originate during female meiosis, and albeit positively correlated with maternal age, age alone is not always predictive of the risk of generating an aneuploid embryo. Therefore, gene variants might account for incorrect chromosome segregation during oogenesis. Given that access to human oocytes is limited for research purposes, a series of assays were developed to study human gene function during meiosis I using mouse oocytes. First, messenger RNA (mRNA) of the gene and gene variant of interest are microinjected into prophase I-arrested mouse oocytes. After allowing time for expression, oocytes are synchronously released into meiotic maturation to complete meiosis I. By tagging the mRNA with a sequence of a fluorescent reporter, such as green fluorescent protein (Gfp), the localization of the human protein can be assessed in addition to the phenotypic alterations. For example, gain or loss of function can be investigated by establishing experimental conditions that challenge the gene product to fix meiotic errors. Although this system is advantageous in investigating human protein function during oogenesis, adequate interpretation of results should be undertaken given that protein expression is not at endogenous levels and, unless controlled for (i.e. knocked out or down), murine homologs are also present in the system.

As the genetic variants associated with human disease begin to become uncovered, it is becoming increasingly important to develop systems with which to rapidly evaluate the biological significance of those identified variants. This protocol describes methods for evaluating human gene function during female meiosis I using mouse oocytes.

マウス卵母細胞を減数分裂の間に人間の遺伝子の機能を評価するために使用私

Embryonic aneuploidy is the major genetic cause of infertility in humans. Most of these events originate during female meiosis, and albeit positively ...

遺伝学

Use of Single Molecule Fluorescent In Situ Hybridization (SM-FISH) to Quantify and Localize mRNAs in Murine Oocytes

Current methods routinely used to quantify mRNA in oocytes and embryos include digital reverse-transcription polymerase chain reaction (dPCR), quantitative, real-time RT-PCR (RT-qPCR) and RNA sequencing. When these techniques are performed using a single oocyte or embryo, low-copy mRNAs are not reliably detected. To overcome this problem, oocytes or embryos can be pooled together for analysis; however, this often leads to high variability amongst samples. In this protocol, we describe the use of fluorescence in situ hybridization (FISH) using branched DNA chemistry. This technique identifies the spatial pattern of mRNAs in individual cells. When the technique is coupled with Spot Finding and Tracking&#160;computer software, the abundance of mRNAs in the cell can also be quantified. Using this technique, there is reduced variability within an experimental group and fewer oocytes and embryos are required to detect significant differences between experimental groups. Commercially available branched-DNA SM-FISH kits have been optimized to detect mRNAs in sectioned tissues or adherent cells on slides. However, oocytes do not effectively adhere to slides and some reagents in the kit were too harsh resulting in oocyte lysis. To prevent this lysis, several modifications were made to the FISH kit. Specifically, oocyte permeabilization and wash&#160;buffers designed for the immunofluorescence of oocytes and embryos replaced the proprietary buffers. The permeabilization, washes, and incubations with probes and amplifier were performed in 6-well plates and oocytes were placed on slides at the end of the protocol using mounting media. These modifications were able to overcome the limitations of the commercially available kit, in particular, the oocyte lysis. To accurately and reproducibly count the number of mRNAs in individual oocytes, computer software was used. Together, this protocol represents an alternative to PCR and sequencing to compare the expression of specific transcripts in single cells.

Use of Single Molecule Fluorescent In Situ Hybridization SM-FISH to Quantify and Localize mRNAs in Murine Oocytes

To reproducibly count the numbers of mRNAs in individual oocytes, single molecule RNA fluorescence in situ hybridization (RNA-FISH) was optimized for non-adherent cells. Oocytes were collected, hybridized with the transcript specific probes, and quantified using an image quantification software.

マウス卵母細胞における定量化するため、Localize Mrna に単一分子蛍光そのままの交配 (SM 魚) の使用

Current methods routinely used to quantify mRNA in oocytes and embryos include digital reverse-transcription polymerase chain reaction (dPCR), ...

Microinjection Techniques for Studying Mitosis in the Drosophila melanogaster Syncytial Embryo

This protocol describes the use of the Drosophila melanogaster syncytial embryo for studying mitosis1. Drosophila has useful genetics with a sequenced genome, and it can be easily maintained and manipulated. Many mitotic mutants exist, and transgenic flies expressing functional fluorescently (e.g. GFP) - tagged mitotic proteins have been and are being generated. Targeted gene expression is possible using the GAL4/UAS system2. 

The Drosophila early embryo carries out multiple mitoses very rapidly (cell cycle duration, &asymp;10 min). It is well suited for imaging mitosis, because during cycles 10-13, nuclei divide rapidly and synchronously without intervening cytokinesis at the surface of the embryo in a single monolayer just underneath the cortex. These rapidly dividing nuclei probably use the same mitotic machinery as other cells, but they are optimized for speed; the checkpoint is generally believed to not be stringent, allowing the study of mitotic proteins whose absence would cause cell cycle arrest in cells with a strong checkpoint. Embryos expressing GFP labeled proteins or microinjected with fluorescently labeled proteins can be easily imaged to follow live dynamics (Fig. 1). In addition, embryos can be microinjected with function-blocking antibodies or inhibitors of specific proteins to study the effect of the loss or perturbation of their function3. These reagents can diffuse throughout the embryo, reaching many spindles to produce a gradient of concentration of inhibitor, which in turn results in a gradient of defects comparable to an allelic series of mutants. Ideally, if the target protein is fluorescently labeled, the gradient of inhibition can be directly visualized4. It is assumed that the strongest phenotype is comparable to the null phenotype, although it is hard to formally exclude the possibility that the antibodies may have dominant effects in rare instances, so rigorous controls and cautious interpretation must be applied. Further away from the injection site, protein function is only partially lost allowing other functions of the target protein to become evident.

Microinjection Techniques for Studying Mitosis in the Drosophila melanogaster Syncytial Embryo

This protocol describes the use of microinjection and high resolution imaging in the Drosophila melanogaster syncytial embryo to study mitosis.

に有糸分裂研究のためのマイクロインジェクション技術キイロショウジョウバエ合胞体胚

This protocol describes the use of the Drosophila melanogaster syncytial embryo  for studying mitosis1.  Drosophila has useful genetics with a sequenced ...

生物学

Nuclear Migration in the Drosophila Oocyte

Live cell imaging is particularly necessary to understand the cellular and molecular mechanisms that regulate organelle movements, cytoskeleton rearrangements, or polarity patterning within the cells. When studying oocyte nucleus positioning, live-imaging techniques are essential to capture the dynamic events of this process. The Drosophila egg chamber is a multicellular structure and an excellent model system to study this phenomenon because of its large size and availability of numerous genetic tools. During Drosophila mid-oogenesis, the nucleus migrates from a central position within the oocyte to adopt an asymmetric position mediated by microtubule-generated forces. This migration and positioning of the nucleus are necessary to determine the polarity axes of the embryo and the subsequent adult fly. One characteristic of this migration is that it occurs in three dimensions (3D), creating a necessity for live imaging. Thus, to study the mechanisms that regulate nuclear migration, we have developed a protocol to culture the dissected egg chambers and perform live imaging for 12 h by time-lapse acquisitions using spinning-disk confocal microscopy. Overall, our conditions allow us to preserve Drosophila egg chambers alive for a long period of time, thereby enabling the completion of nuclear migration to be visualized in a large number of samples in 3D.

Nuclear Migration in the Drosophila Oocyte

In Drosophila, the oocyte nucleus undergoes microtubule-dependent migration during oogenesis. Here, we describe a protocol that was developed to follow the migration by performing live imaging on egg chambers ex-vivo. Our procedure maintains egg chambers alive for 12 h to acquire multi-position 3D time-lapse movies using spinning-disk confocal microscopy.

ショ ウジョウバエ・ ウーサイトにおける核移動

Live cell imaging is particularly necessary to understand the cellular and molecular mechanisms that regulate organelle movements, cytoskeleton ...

発生生物学

Confocal and Super-Resolution Imaging of Polarized Intracellular Trafficking and Secretion of Basement Membrane Proteins During Drosophila Oogenesis

The basement membrane (BM) - a specialized sheet of extracellular matrix present at the basal side of epithelial cells - is critical for the establishment and maintenance of epithelial tissue morphology and organ morphogenesis. Moreover, the BM is essential for tissue modeling, serving as a signaling platform, and providing external forces to shape tissues and organs. Despite the many important roles that the BM plays during normal development and pathological conditions, the biological pathways controlling the intracellular trafficking of BM-containing vesicles and how basal secretion leads to the polarized deposition of BM proteins are poorly understood. The follicular epithelium of the Drosophila ovary is an excellent model system to study the basal deposition of BM membrane proteins, as it produces and secretes all major components of the BM. Confocal and super-resolution imaging combined with image processing in fixed tissues allows for the identification and characterization of cellular factors specifically involved in the intracellular trafficking and deposition of BM proteins. This article presents a detailed protocol for staining and imaging BM-containing vesicles and deposited BM using endogenously tagged proteins in the follicular epithelium of the Drosophila ovary. This protocol can be applied to address both qualitative and quantitative questions and it was developed to accommodate high-throughput screening, allowing for the rapid and efficient identification of factors involved in the polarized intracellular trafficking and secretion of vesicles during epithelial tissue development.

Confocal and Super-Resolution Imaging of Polarized Intracellular Trafficking and Secretion of Basement Membrane Proteins During Drosophila Oogenesis

The basement membrane is essential for tissue and organ morphogenesis during development. To better understand the mechanisms leading to proper placement of this structure, the protocol presented describes methods to visualize and characterize the intracellular trafficking and secretion of basement membrane proteins in epithelial cells using confocal and super-resolution microscopy.

ショウジョウバエの卵子形成期における分極細胞内輸送と基底膜タンパク質の分泌の共焦点および超解像イメージング

The basement membrane (BM) - a specialized sheet of extracellular matrix present at the basal side of epithelial cells - is critical for the establishment ...

Studying Mitotic Checkpoint by Illustrating Dynamic Kinetochore Protein Behavior and Chromosome Motion in Living Drosophila Syncytial Embryos

The spindle assembly checkpoint (SAC) mechanism is an active signal, which monitors the interaction between chromosome kinetochores and spindle microtubules to prevent anaphase onset until the chromosomes are properly connected. Cells use this mechanism to prevent aneuploidy or genomic instability, and hence cancers and other human diseases like birth defects and Alzheimer's1. A number of the SAC components such as Mad1, Mad2, Bub1, BubR1, Bub3, Mps1, Zw10, Rod and Aurora B kinase have been identified and they are all kinetochore dynamic proteins2. Evidence suggests that the kinetochore is where the SAC signal is initiated. The SAC prime regulatory target is Cdc20. Cdc20 is one of the essential APC/C (Anaphase Promoting Complex or Cyclosome) activators3 and is also a kinetochore dynamic protein4-6. When activated, the SAC inhibits the activity of the APC/C to prevent the destruction of two key substrates, cyclin B and securin, thereby preventing the metaphase to anaphase transition7,8. Exactly how the SAC signal is initiated and assembled on the kinetochores and relayed onto the APC/C to inhibit its function still remains elusive.
Drosophila is an extremely tractable experimental system; a much simpler and better-understood organism compared to the human but one that shares fundamental processes in common. It is, perhaps, one of the best organisms to use for bio-imaging studies in living cells, especially for visualization of the mitotic events in space and time, as the early embryo goes through 13 rapid nuclear division cycles synchronously (8-10 minutes for each cycle at 25 &deg;C) and gradually organizes the nuclei in a single monolayer just underneath the cortex9.
Here, I present a bio-imaging method using transgenic Drosophila expressing GFP (Green Fluorescent Protein) or its variant-targeted proteins of interest and a Leica TCS SP2 confocal laser scanning microscope system to study the SAC function in flies, by showing images of GFP fusion proteins of some of the SAC components, Cdc20 and Mad2, as the example.

Studying Mitotic Checkpoint by Illustrating Dynamic Kinetochore Protein Behavior and Chromosome Motion in Living Drosophila Syncytial Embryos

The kinetochore is where the SAC initiates its signal monitoring the mitotic segregation of the sister chromatids. A method is described to visualize the recruitment and turnover of one of the kinetochore proteins and its coordination with the chromosome motion in Drosophila embryos using a Leica laser scanning confocal system.

リビングでの動的動原体タンパク質の挙動と染色体の動きを図解して有糸分裂チェックポイントを勉強<em&gt;ショウジョウバエ</em&gt;合胞体胚

The spindle assembly checkpoint (SAC) mechanism is an active signal, which monitors the interaction between chromosome kinetochores and spindle ...

Live Imaging of GFP-labeled Proteins in Drosophila Oocytes

The Drosophila oocyte has been established as a versatile system for investigating fundamental questions such as cytoskeletal function, cell organization, and organelle structure and function. The availability of various GFP-tagged proteins means that many cellular processes can be monitored in living cells over the course of minutes or hours, and using this technique, processes such as RNP transport, epithelial morphogenesis, and tissue remodeling have been described in great detail in Drosophila oocytes1,2.
The ability to perform video imaging combined with a rich repertoire of mutants allows an enormous variety of genes and processes to be examined in incredible detail. One such example is the process of ooplasmic streaming, which initiates at mid-oogenesis3,4. This vigorous movement of cytoplasmic vesicles is microtubule and kinesin-dependent5 and provides a useful system for investigating cytoskeleton function at these stages.
Here I present a protocol for time lapse imaging of living oocytes using virtually any confocal microscopy setup.

Live Imaging of GFP-labeled Proteins in Drosophila Oocytes

A protocol for live imaging of GFP-tagged proteins or autofluorescent structures in individual Drosophila oocytes is described.

でGFP標識タンパク質のイメージングを生きる<em&gt;ショウジョウバエ</em&gt;卵母細胞

The  Drosophila oocyte has been established as a versatile system for investigating  fundamental questions such as cytoskeletal function, cell ...

Whole Mount RNA Fluorescent in situ Hybridization of Drosophila Embryos

Assessing the expression pattern of a gene, as well as the subcellular localization properties of its transcribed RNA, are key features for understanding its biological function during development. RNA in situ hybridization (RNA-ISH) is a powerful method used for visualizing RNA distribution properties, be it at the organismal, cellular or subcellular levels 1. RNA-ISH is based on the hybridization of a labeled nucleic acid probe (e.g. antisense RNA, oligonucleotides) complementary to the sequence of an mRNA or a non-coding RNA target of interest 2. As the procedure requires primary sequence information alone to generate sequence-specific probes, it can be universally applied to a broad range of organisms and tissue specimens 3. Indeed, a number of large-scale ISH studies have been implemented to document gene expression and RNA localization dynamics in various model organisms, which has led to the establishment of important community resources 4-11. While a variety of probe labeling and detection strategies have been developed over the years, the combined usage of fluorescently-labeled detection reagents and enzymatic signal amplification steps offer significant enhancements in the sensitivity and resolution of the procedure 12. Here, we describe an optimized fluorescent in situ hybridization method (FISH) employing tyramide signal amplification (TSA) to visualize RNA expression and localization dynamics in staged Drosophila embryos. The procedure is carried out in 96-well PCR plate format, which greatly facilitates the simultaneous processing of large numbers of samples.

Whole Mount RNA Fluorescent in situ Hybridization of Drosophila Embryos

Here we describe a whole-mount fluorescent in situ hybridization (FISH) protocol for determining the expression and localization properties of RNAs expressed during embryogenesis in the fruit fly, Drosophila melanogaster.

ホールマウントRNAを蛍光<em&gt;その場で</emの&gt;ハイブリダイゼーション<em&gt;ショウジョウバエ</em&gt;胚

Assessing  the expression pattern of a gene, as well as the subcellular localization  properties of its transcribed RNA, are key features for ...

Techniques for Imaging Prometaphase and Metaphase of Meiosis I in Fixed Drosophila Oocytes

Chromosome segregation in human oocytes is error prone, resulting in aneuploidy, which is the leading genetic cause of miscarriage and birth defects. The study of chromosome behavior in oocytes from model organisms holds much promise to uncover the molecular basis of the susceptibility of human oocytes to aneuploidy. Drosophila melanogaster is amenable to genetic manipulation, with over 100 years of research, community, and technique development. Visualizing chromosome behavior and spindle assembly in Drosophila oocytes has particular challenges, however, due primarily to the presence of membranes surrounding the oocyte that are impenetrable to antibodies. We describe here protocols for the collection, preparation, and imaging of meiosis I spindle assembly and chromosome behavior in Drosophila oocytes, which allow the molecular dissection of chromosome segregation in this important model organism.

Techniques for Imaging Prometaphase and Metaphase of Meiosis I in Fixed Drosophila Oocytes

We present protocols for the collection, preparation, and imaging of mature Drosophila oocytes. These methods allow the visualization of chromosome behavior and spindle assembly and function during meiosis.

固定における減数分裂Iのイメージング前中期および中期のための技術<em&gt;ショウジョウバエ</em&gt;卵母細胞

Chromosome segregation in human oocytes is error prone, resulting in aneuploidy, which is the leading genetic cause of miscarriage and birth defects. The ...

Visualizing and Tracking Endogenous mRNAs in Live Drosophila melanogaster Egg Chambers

Fluorescence-based imaging techniques, in combination with developments in light microscopy, have revolutionized how cell biologists conduct live cell imaging studies. Methods for detecting RNAs have expanded greatly since seminal studies linked site-specific mRNA localization to gene expression regulation. Dynamic mRNA processes can now be visualized via approaches that detect mRNAs, coupled with microscopy set-ups that are fast enough to capture the dynamic range of molecular behavior. The molecular beacon technology is a hybridization-based approach capable of direct detection of endogenous transcripts in living cells. Molecular beacons are hairpin-shaped, internally quenched, single-nucleotide discriminating nucleic acid probes, which fluoresce only upon hybridization to a unique target sequence. When coupled with advanced fluorescence microscopy and high-resolution imaging, they enable one to perform spatial and temporal tracking of intracellular movement of mRNAs. Although this technology is the only method capable of detecting endogenous transcripts, cell biologists have not yet fully embraced this technology due to difficulties in designing such probes for live cell imaging. A new software application, PinMol, allows for enhanced and rapid design of probes best suited to efficiently hybridize to mRNA target regions within a living cell. In addition, high-resolution, real-time image acquisition and current, open source image analysis software allow for a refined data output, leading to a finer evaluation of the complexity underlying the dynamic processes involved in the mRNA&#39;s life cycle.
Here we present a comprehensive protocol for designing and delivering molecular beacons into Drosophila melanogaster egg chambers. Direct and highly specific detection and visualization of endogenous maternal mRNAs is performed via spinning disc confocal microscopy. Imaging data is processed and analyzed using object detection and tracking in Icy software to obtain details about the dynamic movement of mRNAs, which are transported and localized to specialized regions within the oocyte.

Visualizing and Tracking Endogenous mRNAs in Live Drosophila melanogaster Egg Chambers

Here, we present a protocol for the visualization, detection, analysis and tracking of endogenous mRNA trafficking in live Drosophila melanogaster egg chamber using molecular beacons, spinning disc confocal microscopy, and open-source analysis software.

ライブショウジョウバエメラノガスター卵室における内因性mRNAの可視化と追跡

Fluorescence-based imaging techniques, in combination with developments in light microscopy, have revolutionized how cell biologists conduct live cell ...

Prometaphase の腕凝集度と中期の状態を監視する蛍光In Situハイブリダイゼーション (魚) を使用して私はショウジョウバエの卵母細胞

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この動画の章

マウス卵母細胞を減数分裂の間に人間の遺伝子の機能を評価するために使用私

マウス卵母細胞における定量化するため、Localize Mrna に単一分子蛍光そのままの交配 (SM 魚) の使用

に有糸分裂研究のためのマイクロインジェクション技術キイロショウジョウバエ合胞体胚

ショウジョウバエ・ウーサイトにおける核移動

ショウジョウバエの卵子形成期における分極細胞内輸送と基底膜タンパク質の分泌の共焦点および超解像イメージング

リビングでの動的動原体タンパク質の挙動と染色体の動きを図解して有糸分裂チェックポイントを勉強

でGFP標識タンパク質のイメージングを生きる

ホールマウントRNAを蛍光

固定における減数分裂Iのイメージング前中期および中期のための技術

ライブショウジョウバエメラノガスター卵室における内因性mRNAの可視化と追跡

Prometaphase の腕凝集度と中期の状態を監視する蛍光In Situハイブリダイゼーション (魚) を使用して私はショウジョウバエの卵母細胞

要約

さらに動画を探す

この動画の章

マウス卵母細胞を減数分裂の間に人間の遺伝子の機能を評価するために使用私

マウス卵母細胞における定量化するため、Localize Mrna に単一分子蛍光そのままの交配 (SM 魚) の使用

に有糸分裂研究のためのマイクロインジェクション技術キイロショウジョウバエ合胞体胚

ショ ウジョウバエ・ ウーサイトにおける核移動

ショウジョウバエの卵子形成期における分極細胞内輸送と基底膜タンパク質の分泌の共焦点および超解像イメージング

リビングでの動的動原体タンパク質の挙動と染色体の動きを図解して有糸分裂チェックポイントを勉強

でGFP標識タンパク質のイメージングを生きる

ホールマウントRNAを蛍光

固定における減数分裂Iのイメージング前中期および中期のための技術

ライブショウジョウバエメラノガスター卵室における内因性mRNAの可視化と追跡

ショウジョウバエ・ウーサイトにおける核移動