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This protocol is useful to obtain clear and repeatable data to know the motor function of a worm by counting the number of body thrash in M9 buffer. The thrashing assay is useful for observation of the effect of loss of motor neurons such as VA or VB neuron.

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The Method of the Body Bending Assay Using Caenorhabditis elegans
使用秀丽隐杆线虫进行身体弯曲试验法

神经科学 > 行为神经科学 > 认知
作者: Mikiro Nawa
Mikiro Nawa Affiliation: Department of Pharmacology, Tokyo Medical University, Shinjuku-ku, Tokyo, Japan
Bio-protocol author page: a89
 and Masaaki Matsuoka
Masaaki MatsuokaAffiliation: Department of Pharmacology, Tokyo Medical University, Shinjuku-ku, Tokyo, Japan
For correspondence: sakimatu@tokyo-med.ac.jp
Bio-protocol author page: a90
Vol 2, Iss 17, 9/5/2012, 6874 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.253

[Abstract] This protocol is useful to obtain clear and repeatable data to know the motor function of a worm by counting the number of body thrash in M9 buffer. The thrashing assay is useful for observation of the effect of loss of motor neurons such as VA or VB neuron.

[Abstract] 该实验方案的中文版正在准备中...

Materials and Reagents

  1. KH2PO4 (Wako Chemicals USA, catalog number:  169-04245 )
  2. Na2HPO4 (Wako Chemicals USA, catalog number:  197-02865 )
  3. NaCl (Wako Chemicals USA, catalog number:  191-01665 )
  4. M9 buffer (see Recipes)

Equipment

  1. Sterile NGM agar plate with a 35 mm diameter

Procedure

Fill sterile NGM agar plates with a 35 mm diameter with 1 ml of M9 buffer. Before the start of this assay, put a worm on a sterile NGM agar plate without bacteria and allow it to crawl freely to remove the agglomerated bacteria from the worm. After examining whether the bacteria were removed by visually, put a worm (4-day-old) into the buffer and allow it to swim freely for 1 min to be accustomed to the environment. 
Then count the number of thrash for 1 min (Nawa et al., 2012). Count at least 10 worms for each assay.
Note: A movement of the worm that swings its head and/or tail to the same side is counted as one thrash. A movie of body-bending worms makes counting more correct and easy. The bacteria that are served as food for worms should be removed as completely as possible because they affect the worm's movement.

Recipes

  1. M9 buffer (Brenner, 1974)

Acknowledgments

This protocol is adapted from Brenner (1974) and previously used in Nawa et al. (2012).

References

  1. Brenner, S. (1974). The genetics of Caenorhabditis elegans. Genetics 77(1): 71-94.
  2. Nawa, M., Kage-Nakadai, E., Aiso, S., Okamoto, K., Mitani, S. and Matsuoka, M. (2012). Reduced expression of BTBD10, an Akt activator, leads to motor neuron death. Cell Death Differ 19(8): 1398-1407.

材料和试剂

  1. KH sub 2 PO 4(Wako Chemicals USA,目录号:169-04245)
  2. Na 2 HPO 4(Wako Chemicals USA,目录号:197-02865)
  3. NaCl(Wako Chemicals USA,目录号:191-01665)
  4. M9缓冲区(请参阅配方)

设备

  1. 具有35mm直径的无菌NGM琼脂板

程序

填充35毫米直径与1毫升M9缓冲液的无菌NGM琼脂板。在开始这种测定之前,将蠕虫放在无细菌的无菌NGM琼脂板上,并允许其自由爬行以从蠕虫中除去附聚的细菌。在检查细菌是否通过视觉去除后,将蠕虫(4天龄)放入缓冲液中,并允许其自由游动1分钟以习惯环境。
然后计数1分钟的瘙痒次数(Nawa等人,2012)。每个测定计数至少10个蠕虫。
注意:将其头部和/或尾部摆动到同一侧的蠕虫的运动被计为一个鞭毛。身体弯曲蠕虫的电影使计数更正确和容易。用作蠕虫食物的细菌应该尽可能完全地去除,因为它们会影响蠕虫的运动。

食谱

  1. M9缓冲区(Brenner,1974)

致谢

该协议改编自Brenner(1974)并且之前在Nawa等人(2012)中使用。

参考文献

  1. Brenner,S.(1974)。 Caenorhabditis elegans的遗传学 遗传学 77(1):71-94。
  2. Nawa,M.,Kage-Nakadai,E.,Aiso,S.,Okamoto,K.,Mitani,S。和Matsuoka,M。(2012)。 减少BTBD10(一种Akt激活剂)的表达,导致运动神经元死亡。 Cell Death Differ 19(8):1398-1407。
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How to cite this protocol: Nawa, M. and Matsuoka, M. (2012). The Method of the Body Bending Assay Using Caenorhabditis elegans. Bio-protocol 2(17): e253. DOI: 10.21769/BioProtoc.253; Full Text



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