娜姐朱佰利群同等贡献这项工作。这项工作在财政的公益专项基金林业科学研究（201204501）和中国国家自然科学基金（31170604，31270688，31570638和）的支持。我们感谢Holighaus格里特博士和Danielle Hickford博士对英语写作的有益建议。

1.处女成人线虫收购<ol><li> 松材线虫分离和培养条件 注: 松材线虫隔离Nxy61最初是从浙江省在中国的宁波地区患病马尾松提取。 <ol><li>准备马铃薯葡萄糖琼脂（PDA，见材料表 ）种植灰霉病板。 </li><li>对于接种，冲了一块与从灰霉病垫子10毫米直径的真菌垫，并把它放在一个新的PDA板中间。文化在黑暗中灰霉病 5天在25℃。 </li><li>文化包含灰霉病在25°C黑暗准备的培养皿松木线虫。 </li></ol></li><li> 松材线虫鸡蛋收购 <ol><li>将一个漏斗下方的夹紧橡胶管按住准备漏斗在机架组装贝尔曼漏斗装置。放置滤纸代替两层纱布在漏斗口。 </li><li>转移真菌培养漏斗设置，直到真菌垫浸入加水。等待2小时，线虫会从真菌培养出来，穿过滤纸，并在贝尔曼漏斗的最低部分安顿下来。 </li><li>从管的底部成15毫升管收集第一对的水滴，并通过缓慢释放在管夹紧收集线虫。 </li><li>离心上述集合以3000 xg离心在RT 2分钟。 </li><li>用移液管除去水的上层，并用1毫升无菌双蒸（DD）的H 2 O重悬它</li><li>用移液管，在上述再悬浮转移至新的玻璃培养皿（直径6cm）。加入3毫升无菌DD 的 H 2 O至培养皿以确保线虫能够自由游泳。 </li><li>保持线虫在GLASš的Petri在黑暗盘10分钟在25℃。鸡蛋会被怀孕女性被解雇。因为它们的糖蛋白表面的，将卵粘在玻璃培养皿的底部。 </li><li>小心除去水和蠕虫，而不会干扰蛋。加入3毫升无菌DD 的 H 2 O至玻璃培养皿立即防止蛋干燥。 </li><li>重复步骤3倍1.2.8删除所有的幼虫和成虫，并获得纯净的鸡蛋。添加无菌DD 的 H 2的O的额外2毫升（5毫升计）的培养皿。 </li></ol></li><li> 第四阶段少年的采集（J4的幼虫） <ol><li>哈奇收集鸡蛋在黑暗中24小时在25℃，得到J2幼虫。 </li><li>用移液管转移J2幼虫到15毫升管中，离心以3000 xg离心2分钟，并除去水的上层。 </li><li>重悬用无菌DD H 2的O 200微升J2幼虫，然后将它们转移到含灰霉病的PDA板。 </li><li>文化在黑暗中J2幼虫52 h后25℃。在这个时间点，最J2幼虫会已发展成J4幼虫的早期阶段。 </li></ol></li><li> 处女的成年线虫的收购 <ol><li>准备通过在酒精灯加热后绘制的玻璃毛细管拿起线虫针。 注:此毛细管针的尖端通常需要为50微米的直径，大致成年线虫的主体的宽度，因此，它可以用作用于拾取单个线虫的工具。 </li><li>组装贝尔曼漏斗并提取从真菌培养物线虫2小时，如在步骤1.2.1描述 - 1.2.3。 </li><li>传输与水J4到一个干净的培养皿。 </li><li>观察在立体显微镜下的线虫和生殖器形态学男性和女性之间区分，如通过玛米亚和清原8（ 图1中描述</strong&gt;的）。拾取所需性别的线虫与制备毛细管针，并将其转移到合适的真菌培养板为男性或女性。 注:有女性多于男性，而女性在60百分比 - 70％。女性外阴是从男性毛刺明显不同。相比于成人，在早期J4的毛刺或外阴更小，但它们仍然可以很容易地看到和分辨。线虫总是在水中，这有时会增加在区分男性和女性的困难移动。 </li><li>培养J4的幼虫在暗处在25℃另外24小时。 注:所有线虫应发育为成虫，但因为它们是未交配，它们被认为是处女成年人。 </li><li>装配2贝尔曼漏斗和提取的真菌培养处女大人2小时，如步骤1.2.1描述 - 1.2.3，和处女的成年男性和女性转移到两个独立的，清洁的培养皿。 </li></o升&gt; </li></ol> 2.观察和交配行为的录像<ol><li>准备几个凹幻灯片，并添加无菌DD H 2的O 200μL到他们的每一个孔中。 </li><li>拿起准备好的处女的成年男性或女性，并用毛细管针，把只有一个线虫到每个凹幻灯片的水。 </li><li>保持井内的处女大人1小时，让他们适应新的水环境中。 </li><li>用吸管转移其中的一个从一个井到另一对处女的男性和女性。 </li><li>观察立体显微镜下的凹幻灯片1♀+1♂组合的交配行为。 </li><li>分析处女成年人的配合效率，计算出它们的交配成功率（R）和其与随机选择的成人对的比较。计算R相比研究重复的总数，其中观察到交配重复的百分比。 </li><li&gt;探讨交配选择和竞争性内，组和观察两种不同的组合处女线虫:1♀+3♂和3♀+1♂。 </li><li>为了收集数据进行定性和定量分析，电影利用视频显微镜的交配行为。通常情况下，整个交配过程，从时间，当线虫放置在凹幻灯片一时的好，当他们交配后搬走时，2小时内完成。因此，连续地记录了至少2小时作为一个复制的配合过程。完成至少30复制总数为每个组合。 注:添加更多无菌DD H 2 O（通常为50微升）每0.5小时至井的录制过程中，以弥补蒸发。 </li></ol> 3.数据收集<ol><li>仔细查看电脑屏幕上的所有视频和定性描述子的行为。 </li><li>通过测量次交配的时间Ë帧一帧的视频录像机功能。手动减去从结束点的开始时间。 </li><li>对于1♀+1♂组合，分析每一个视频录制和测量以下事件的持续时间: <ol><li>测量搜索的持续时间（ 即，从当阴和阳被引入凹形滑动到它们遇到的那一刻的阱和彼此接触的时间）。 </li><li>测量接触的持续时间（ 即 ，从第一接触的瞬间当阳被精确地定位在阴的外阴用于第一交配的时间）。 </li><li>测量交尾的持续时间（ 即 ，从当直到他射精后滑动从女性的身体向下阳被精确地定位在阴的外阴的时间）。注意:某些线虫可在2小时交配几次。在这种情况下，测量交配从第一交配期间向拉斯的端t的一，包括之间的时间。 </li><li>测量挥之不去的持续时间（ 即 ，交配后的期间，直到彼此雌雄游程，通过的距离超过所述线虫的身体长度被分离（通常1毫米））。 </li></ol></li><li>计数交配次数每交配的持续时间和所有交配的在2小时的平均持续时间为每个不同的组合（ 即，1♀+1♂，3♀+1♂和1♀+3♂）。 </li><li>计算第一接触（P），对于不同的组合作为第一接触，这是立即和没有任何中断进行后交配成功率的交配成功率。在接触阶段，几次接触可能实际交配后方可。 <ol><li>当两个个体之间的距离小于所述主体的长度以上，考虑一个接触到结束。对于一个复制，记录数p作为100％时，交配我小号马上第一次接触后，如果没有观察到交配或观察到的，没有休息，或0.00％，如果有交配但经过两个或两个以上的联系人。 注:有30个重复这里描述的每个组合。 </li></ol></li></ol>

<ol>
	<li>Tokushige, Y., Kiyohara, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bursaphelenchus+sp.+in+the+wood+of+dead+pine+trees.">Bursaphelenchus sp. in the wood of dead pine trees.</a> J. Jap. For. Soc. 51, 193-195 (1969).</li><li>Sun, Y. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bursaphelenchus+xylophilus+was+discovered+in+Sun+Yet-sen's+mausoleum+in+Nanjing.">Bursaphelenchus xylophilus was discovered in Sun Yet-sen's mausoleum in Nanjing.</a> J. Jiangsu Fore. Sci. Tech. (in Chinese). 4, 47 (1982).</li><li>Mota, M. 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=First+report+of+Bursaphelenchus+xylophilus+in+Portugal+and+in+Europe.">First report of Bursaphelenchus xylophilus in Portugal and in Europe.</a> Nematology. 1, 727-734 (1999).</li><li>Robertson, 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=Incidence+of+the+pine+wood+nematode+Bursaphelenchus+xylophlius+Steiner+&amp;+Buhrer,+1934+(Nickle,+1970)+in+Spain.">Incidence of the pine wood nematode Bursaphelenchus xylophlius Steiner &amp; Buhrer, 1934 (Nickle, 1970) in Spain.</a> Nematology. 13, 755-757 (2011).</li><li>Vicente, C., Espada, M., Vieira, P., Mota, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pine+wilt+disease:+a+threat+to+European+forestry.">Pine wilt disease: a threat to European forestry.</a> Euro. J. Plant Path. 133, 89-99 (2012).</li><li>Jones, J. T., Moens, M., Mota, M., Li, H., Kikuchi, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bursaphelenchus+xylophilus:+opportunities+in+comparative+genomics+and+molecular+host-parasite+interactions.">Bursaphelenchus xylophilus: opportunities in comparative genomics and molecular host-parasite interactions.</a> Mol. Plant. Path. 9, 357-368 (2008).</li><li>Mamiya, Y., Furukawa, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fecundity+and+reproduction+of+Bursaphelenchus+lignicolus.">Fecundity and reproduction of Bursaphelenchus lignicolus.</a> Jap. J. Nemat. 7, 6-9 (1977).</li><li>Mamiya, Y., Kiyohara, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Description+of+Bursaphelenchus+lignicolus.+n.+sp.+(Nematoda:+Aphelenchoididae)+from+pine+wood+and+histophathology+of+nematode-infested+trees.">Description of Bursaphelenchus lignicolus. n. sp. (Nematoda: Aphelenchoididae) from pine wood and histophathology of nematode-infested trees.</a> Nematologica. 18, 120-124 (1972).</li><li>Liu, B. J., 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=Behavioural+features+of+Bursaphelenchus+xylophilus+in+the+mating+process.">Behavioural features of Bursaphelenchus xylophilus in the mating process.</a> Nematology. 16, 895-902 (2014).</li><li>Simon, J. M., Sternberg, P. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evidence+of+a+mate+finding+cue+in+the+hermaphrodite+nematode+Caenorhabditis+elegans.">Evidence of a mate finding cue in the hermaphrodite nematode Caenorhabditis elegans.</a> Proc. Natl. Acad. Sci. USA. 99, 1598-1603 (2002).</li><li>Barr, M. M., Garcia, L. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Male+mating+behaviour.">Male mating behaviour.</a> The C. elegans Research Community Wormbook. , (2006).</li><li>Shinya, R., Chen, A., Sternberg, P. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sex+attraction+and+mating+in+Bursaphelenchus+okinawaensis+and+B.+xylophilus.">Sex attraction and mating in Bursaphelenchus okinawaensis and B. xylophilus.</a> J. Nematol. 47 (3), 176-183 (2015).</li></ol>

The authors have nothing to disclose.

行为测试是化学生态学，神经生物学，分子生物学和遗传学的一个基本方面。线虫，特别是线虫 ，被广泛地用于这些领域的研究。 线虫的交配行为之前已10-11调查。但是， 松材线虫是雌雄异体，并从雌雄同体线虫不同，它的交配过程有不同的行为特征9。近日， 松材线虫okinawaensis， 松材线虫的雌雄同体的妹妹种的交配行为报12。...

The pinewood nematode, Bursaphelenchus xylophilus (Steiner and Buhrer) Nickle, is one of the most destructive invasive species, causing the wilting and eventual death of pine trees. This pathogenic nematode is believed to be native to the USA, but it was transported to several countries, including China, Japan, South Korea, and Portugal. Recently, it has also been reported in Spain, killing millions of pine trees at huge economic losses and threatening forest production and ecological stability1-5.
Once a host pine tree is infected by pinewood nematodes, thousands of millions of offspring are propagated rapidly within the trunk. This causes xylem dysfunction, resulting in wilting and eventually in the death of the host tree6. However, there is currently no efficient way to control this disease. Mating behavior might play an important role in the high fecundity of this nematode7. We therefore investigated the mating behavior of B. xylophilus in the laboratory, trying to find an effective way of disturbing its mating and decreasing its fecundity. 
This protocol is intended to introduce the detailed methods of how to obtain the virgin adults of B. xylophilus and how to observe and analyze the mating behavior using a video recorder and a stereomicroscope. This protocol can also be used as a reference for behavioral studies of other nematodes.

<table><tbody><tr><td>&lt;em&gt;B. xylophilus&lt;/em&gt; isolate &lt;em&gt;Nxy61&lt;/em&gt;</td><td></td><td></td><td>Extracted from &lt;em&gt;Pinus massoniana&lt;/em&gt; in Ningbo area of China</td></tr><tr><td>Grey mold strain</td><td></td><td></td><td>Obtained from Forestry Academy of China</td></tr><tr><td>Baermann funnels</td><td>Sengong</td><td></td><td></td></tr><tr><td>falcon tubes</td><td>Sengong</td><td></td><td></td></tr><tr><td>centrifuge</td><td>Sengong</td><td></td><td></td></tr><tr><td>pipette</td><td>Sengong</td><td></td><td></td></tr><tr><td>sterile dd H&lt;sub&gt;2&lt;/sub&gt;O</td><td></td><td></td><td>Sterilized by high pressure of steam at 121 &amp;deg;C for 30 min.</td></tr><tr><td>Petri dishes</td><td>Sengong</td><td></td><td></td></tr><tr><td>PDA medium</td><td>Huankai Bio.</td><td>021050</td><td></td></tr><tr><td>incubator</td><td>Sengong</td><td></td><td></td></tr><tr><td>Laminar flow</td><td>Sengong</td><td></td><td></td></tr><tr><td>glass capillary</td><td>Sengong</td><td></td><td></td></tr><tr><td>Stereoscope</td><td>Leica</td><td>LED5000 RL</td><td></td></tr><tr><td>inverted stereo microscope</td><td>Zeiss</td><td>A1</td><td></td></tr><tr><td>concave slides</td><td>Sengong</td><td></td><td></td></tr></tbody></table>

observation and quantification of mating behavior in the pinewood nematode, bursaphelenchus xylophilus

用于观察和量化的松材线虫， 松材线虫的交配行为的方法，是立体显微镜下成立的。以提高松材线虫的配合效率和增加的交配观测的几率，处女成年人中培养并用于调查。卵通过保持线虫在水中，并允许雌性产卵10分钟获得的。第二级幼体（J2）通过在黑暗中在25℃温育卵24小时同步，和J4早期通过培养用灰霉病， 灰霉病在J2，另一个52 h条件。在这个时间点，最J4线虫可以作为是男性或女性使用其生殖器形态明确区分。男性和女性的J4收集并在两个不同的培养皿中24小时分别培养以获得处女成人线虫。一个处女男和处女女在笏一滴配对器中的凹滑动的孔中。交配行为与立体显微镜下录像机拍摄。配合过程的整个时间为82.8±3.91分钟（平均值±SE）和可分为4个不同的阶段:搜索，接触，交尾，和挥之不去。持续时间的平均分者21.8±2.0，28.0±1.9，25.8±0.7和7.2±0.5，分别为。 11个子行为进行了描述:巡航，接近，遭遇，触摸，胡平，定位，安装，射精，分离，平静，和漫游。有趣的是，当一个女性与男性3个或一个男性，女性3分组观察到明显的内部性的竞争。这个协议是有用的，有价值的，不仅在调查松材线虫的交配行为，而且在充当其他线虫行为学研究的参考。

处女成人的交配成功率（R）为86.67％，平均，比该随机选择的成人显著更高:13.33±4.65％（F = 1301.71，DF = 1，P = 0.0001）（ 图2）。这一发现表明，它更容易观察松材线虫与处女比成年人随机选取成年人交配行为。 当一名男性与一名女性配对，整个交配过程持续了平均82.8分钟。整个过程可以通过四...

Watch this Scientific Journal Video about Observation and Quantification of Mating Behavior in the Pinewood Nematode, Bursaphelenchus xylophilus at JoVE.com

Observation and Quantification of Mating Behavior in the Pinewood Nematode, Bursaphelenchus xylophilus

A protocol for investigating the mating behavior of the pinewood nematode, Bursaphelenchus xylophilus, is presented. Behavioral features of B. xylophilus are described in the mating process.

观察和交配行为的量化在我国松材线虫，<em&gt;松材线虫</em

A method for observing and quantifying the mating behavior of the pinewood nematode, Bursaphelenchus xylophilus, was established under a stereomicroscope. ...

observation-quantification-mating-behavior-pinewood-nematode

Research

JoVE Journal

Biology

Observation and Quantification of Mating Behavior in the Pinewood Nematode, Bursaphelenchus xylophilus

9.4K 次观看.  Zhejiang Agricultural & Forestry University. A protocol for investigating the mating behavior of the pinewood nematode, Bursaphelenchus xylophilus, is presented. Behavioral features of B. xylophilus are described in the mating process.

视频: Observation and Quantification of Mating Behavior in the Pinewood Nematode, Bursaphelenchus xylophilus

Technique for Studying Arthropod and Microbial Communities within Tree Tissues

Phloem tissues of pine are habitats for many thousands of organisms. Arthropods and microbes use phloem and cambium tissues to seek mates, lay eggs, rear young, feed, or hide from natural enemies or harsh environmental conditions outside of the tree. Organisms that persist within the phloem habitat are difficult to observe given their location under bark. We provide a technique to preserve intact phloem and prepare it for experimentation with invertebrates and microorganisms. The apparatus is called a &#8216;phloem sandwich&#8217; and allows for the introduction and observation of arthropods, microbes, and other organisms. This technique has resulted in a better understanding of the feeding behaviors, life-history traits, reproduction, development, and interactions of organisms within tree phloem. The strengths of this technique include the use of inexpensive materials, variability in sandwich size, flexibility to re-open the sandwich or introduce multiple organisms through drilled holes, and the preservation and maintenance of phloem integrity. The phloem sandwich is an excellent educational tool for scientific discovery in both K-12 science courses and university research laboratories.

We provide a technique to preserve intact tree phloem and prepare it for observation. We create an apparatus called a phloem sandwich that allows for the introduction and observation of arthropods, microbes, and other organisms that inhabit phloem tissues.

技术树内组织学习节肢动物和微生物群落

Phloem tissues of pine are habitats for many thousands of organisms.  Arthropods and microbes use phloem and cambium tissues to seek mates, lay eggs, rear ...

科研

环境科学

Assaying Predatory Feeding Behaviors in Pristionchus and Other Nematodes

This protocol provides multiple methods for the analysis and quantification of predatory feeding behaviors in nematodes. Many nematode species including Pristionchus pacificus display complex behaviors, the most striking of which is the predation of other nematode larvae. However, as these behaviors are absent in the model organism Caenorhabditis elegans, they have thus far only recently been described in detail along with the development of reliable behavioral assays 1. These predatory behaviors are dependent upon phenotypically plastic but fixed mouth morphs making the correct identification and categorization of these animals essential. In P. pacificus there are two mouth types, the stenostomatous and eurystomatous morphs 2, with only the wide mouthed eurystomatous containing an extra tooth and being capable of killing other nematode larvae. Through the isolation of an abundance of size matched prey larvae and subsequent exposure to predatory nematodes, assays including both "corpse assays" and "bite assays" on correctly identified mouth morph nematodes are possible. These assays provide a means to rapidly quantify predation success rates and provide a detailed behavioral analysis of individual nematodes engaged in predatory feeding activities. In addition, with the use of a high-speed camera, visualization of changes in pharyngeal activity including tooth and pumping dynamics are also possible.

Assaying Predatory Feeding Behaviors in Pristionchus and Other Nematodes

Behavioral assays provide powerful tools for understanding neuronal function. Here we present several protocols for quantifying predatory feeding behavior found in the model nematode Pristionchus pacificus and its relatives. Additionally, we provide methods for analyzing predatory feeding adaptations including mouth structures and teeth.

测定掠夺取食行为<em&gt; Pristionchus</em&gt;及其他线虫

This protocol provides multiple methods for the analysis and quantification of predatory feeding behaviors in nematodes. Many nematode species including ...

Induction and Evaluation of Inbreeding Crosses Using the Ant, Vollenhovia Emeryi

The genetic and molecular components of the sex-determination cascade have been extensively studied in the honeybee, Apis mellifera, a hymenopteran model organism. However, little is known about the sex-determination mechanisms found in other non-model hymenopteran taxa, such as ants. Because of the complex nature of the life cycles that have evolved in hymenopteran species, it is difficult to maintain and conduct experimental crosses between these organisms in the laboratory. Here, we describe the methods for conducting inbreeding crosses and for evaluating the success of those crosses in ant Vollenhovia emeryi. Inducing inbreeding in the laboratory using V. emeryi, is relatively simple because of the unique biology of the species. Specifically, this species produces androgenetic males, and female reproductives exhibit wing polymorphism, which simplifies identification of the phenotypes in genetic crosses. In addition, evaluating the success of inbreeding is straightforward as males can be produced continuously by inbreeding crosses, while normal males only appear during a well-defined reproductive season in the field. Our protocol allow for using V. emeryi as a model to investigate the genetic and molecular basis of the sex determination system in ant species.

Induction and Evaluation of Inbreeding Crosses Using the Ant, Vollenhovia Emeryi

In this protocol, methods for conducting inbreeding crosses, and for evaluating the success of those crosses, are described for the ant Vollenhovia emeryi. These protocols are important for experiments aimed at understanding the genetic basis of sex determination systems in Hymenoptera.

利用蚂蚁Vollenhovia Emeryi对近亲繁殖杂交的诱导和评价

The genetic and molecular components of the sex-determination cascade have been extensively studied in the honeybee, Apis mellifera, a hymenopteran model ...

遗传学

A Method to Test the Effect of Environmental Cues on Mating Behavior in Drosophila melanogaster

An individual's sexual drive is influenced by genotype, experience and environmental conditions. How these factors interact to modulate sexual behaviors remains poorly understood. In Drosophila melanogaster, environmental cues, such as food availability, affect mating activity offering a tractable system to investigate the mechanisms modulating sexual behavior. In D. melanogaster, environmental cues are often sensed via the chemosensory gustatory and olfactory systems. Here, we present a method to test the effect of environmental chemical cues on mating behavior. The assay consists of a small mating arena containing food medium and a mating couple. The mating frequency for each couple is continuously monitored for 24 h. Here we present the applicability of this assay to test environmental compounds from an external source through a pressurized air system as well as manipulation of the environmental components directly in the mating arena. The use of a pressurized air system is especially useful to test the effect of very volatile compounds, while manipulating components directly in the mating arena can be of value to ascertain a compound's presence. This assay can be adapted to answer questions about the influence of genetic and environmental cues on mating behavior and fecundity as well as other male and female reproductive behaviors.

A Method to Test the Effect of Environmental Cues on Mating Behavior in Drosophila melanogaster

We demonstrate an assay to analyze the environmental and genetic cues that influence mating behavior in the fruit fly Drosophila melanogaster.

测试环境线索对交配行为的影响的方法<em&gt;黑腹果蝇</em

An individual's sexual drive is influenced by genotype, experience and environmental conditions. How these factors interact to modulate sexual behaviors ...

行为学

Individual Culturing of Tigriopus Copepods and Quantitative Analysis of Their Mate-guarding Behavior

Copepods of the genus Tigriopus, which are common zooplankton in rocky tide pools, show precopulatory mate-guarding behavior where a male clasps a potential mate to form a pair. While this phenomenon has attracted interest of researchers, methods for its analysis have not been well described. Here we describe procedures for: 1) individual culturing and staging of Tigriopus juveniles and adults, and 2) video-based analysis of their mate-guarding behavior. The culturing method enables experimental control of paring experience of animals as well as the ability to track their development before behavioral tests. The analysis method allows quantitative evaluation of several aspects of the mate-guarding behavior, including capturing attempts by males and swimming trajectory of mate-guarding pairs. Although these methods were originally established for ethological studies on Tigriopus, with proper modifications they can also be applied to studies of other zooplankton in different research fields, such as physiology, toxicology, and ecological genetics.

Individual Culturing of Tigriopus Copepods and Quantitative Analysis of Their Mate-guarding Behavior

Mate-guarding behavior plays an important role in reproduction of intertidal copepods of the genus Tigriopus. However, methods for studying this behavior have not been well described. Here we describe methods for: 1) individual culture of virgin Tigriopus animals, and 2) quantitative analysis of their mate-guarding behavior.

Tigriopus桡足的个体培养及其交配行为的定量分析

Copepods of the genus Tigriopus, which are common zooplankton in rocky tide pools, show precopulatory mate-guarding behavior where a male clasps a ...

Rapid Isolation of Wild Nematodes by Baermann Funnel

Beyond being robust experimental model organisms, Caenorhabditis elegans and its relatives are also real animals that live in nature. Studies of wild nematodes in their natural environments are valuable for understanding many aspects of biology, including the selective regimes in which distinctive genomic and phenotypic characters evolve, the genetic basis for complex trait variation, and the natural genetic diversity fundamental to all animal populations. This manuscript describes a simple and efficient method for extracting nematodes from their natural substrates, including rotting fruits, flowers, fungi, leaf litter, and soil. The Baermann funnel method, a classical nematology technique, selectively isolates active nematodes from their substrates. Because it recovers nearly all active worms from the sample, the Baermann funnel technique allows for the recovery of rare and slow-growing genotypes that co-occur with abundant and fast-growing genotypes, which might be missed in extraction methods that involve multiple generations of reproduction. The technique is also well suited to addressing metagenetic, population-genetic, and ecological questions. It captures the entire population in a sample simultaneously, allowing an unbiased view of the natural distribution of ages, sexes, and genotypes. The protocol allows for deployment at scale in the field, rapidly converting substrates into worm plates, and the authors have validated it through fieldwork on multiple continents.

This protocol outlines a method for efficiently extracting live nematodes from natural substrates in the field.

由Baermann Funnel快速分离野生线虫

Beyond being robust experimental model organisms, Caenorhabditis elegans and its relatives are also real animals that live in nature. Studies of wild ...

<meta charset="utf8"></head><body>秀丽隐杆线虫中挥发性信息素反应的提取和定量

Volatile Sex Pheromone Extraction and Chemoattraction Assay in Caenorhabditis elegans

Chemical communication is vital in organismal health, reproduction, and overall well-being. Understanding the molecular pathways, neural processes, and computations governing these signals remains an active area of research. The nematode Caenorhabditis elegans provides a powerful model for studying these processes as it produces a volatile sex pheromone. This pheromone is synthesized by virgin females or sperm-depleted hermaphrodites and serves as an attractant for males.
This protocol describes a detailed method for isolating the volatile sex pheromone from several C. elegans strains (WT strain N2, daf-22, and fog-2) and C. remanei. We also provide a protocol for quantifying the male chemotaxis response to the volatile sex pheromone. Our analysis utilizes measurements such as chemotaxis index (C.I.), arrival time (A.T.), and a trajectory plot to compare male responses under various conditions accurately. This method allows for robust comparisons between males of different genetic backgrounds or developmental stages. Furthermore, the experimental setup outlined here is adaptable to investigating other chemoattraction chemicals.

<meta charset="utf8"></head><body>作者聚焦：检查挥发性信息素对雄性 秀丽隐杆线 虫行为的影响

This protocol establishes methods for extracting and quantifying responses to the volatile sex pheromone in C. elegans, providing tools to study chemical communication and navigation trajectory.

秀丽隐杆线虫中的挥发性信息素提取和趋化试验

Chemical communication is vital in organismal health, reproduction, and overall well-being. Understanding the molecular pathways, neural processes, and ...

Measuring and Altering Mating Drive in Male Drosophila melanogaster

Despite decades of investigation, the neuronal and molecular bases of motivational states remain mysterious. We have recently developed a novel, reductionist, and scalable system for in-depth investigation of motivation using the mating drive of male Drosophila&#160;melanogaster (Drosophila), the methods for which we detail here. The behavioral paradigm centers on the finding that male mating drive decreases alongside fertility over the course of repeated copulations and recovers over ~3 d. In this system, the powerful neurogenetic tools available in the fly converge with the genetic accessibility and putative wiring diagram available for sexual behavior. This convergence allows rapid isolation and interrogation of small neuronal populations with specific motivational functions. Here we detail the design and execution of the satiety assay that is used to measure and alter courtship motivation in the male fly. Using this assay, we also demonstrate that low male mating drive can be overcome by stimulating dopaminergic neurons. The satiety assay is simple, affordable, and robust to influences of genetic background. We expect the satiety assay to generate many new insights into the neurobiology of motivational states.

Measuring and Altering Mating Drive in Male Drosophila melanogaster

This article describes a behavioral assay that uses male mating drive in Drosophila melanogaster to study motivation. Using this method, researchers can utilize advanced fly neurogenetic techniques to uncover the genetic, molecular, and cellular mechanisms that underlie this motivation.

观察和交配行为的量化在我国松材线虫，

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技术树内组织学习节肢动物和微生物群落

测定掠夺取食行为

利用蚂蚁Vollenhovia Emeryi对近亲繁殖杂交的诱导和评价

测试环境线索对交配行为的影响的方法

Tigriopus桡足的个体培养及其交配行为的定量分析

由Baermann Funnel快速分离野生线虫

秀丽隐杆线虫中的挥发性信息素提取和趋化试验

秀丽隐杆线虫中的挥发性信息素提取和趋化试验

秀丽隐杆线虫中的挥发性信息素提取和趋化试验

测量并改变配套驱动马累