搜索

Locomotor Assay in Drosophila melanogaster
黑腹果蝇的运动分析   

评审
匿名评审
下载 PDF 引用 收藏 提问与回复 分享您的反馈 Cited by

本文章节

Abstract

This protocol describes a simple locomotor assay in Drosophila melanogaster. In brief, the locomotor of each single fly in the culture dish is recorded by a web camera. The moving time, walking length, speed and the locomotor trails of the single fly could be quantitatively analyzed.

Keywords: Locomotor(运动), Drosophila(果蝇), Video analysis(视频分析), Motion detection(运动检测), Motion trail(运动轨迹), Behavior(行为)

Background

This protocol was implemented in the previously published study (Liu et al., 2016). In that study, this assay was combined with the optogenetic system by simply providing the proper excitation light.

Materials and Reagents

  1. 3.5 cm culture dishes with black paper inside
  2. Drosophila strains to be analyzed
  3. Ice for anesthesia

Equipment

  1. Empty vial for cold anesthesia
  2. Fluorescent lamp (Bannet T5 8 W, China)
  3. Infrared LEDs (TUOENS, model: TS-6036A )
  4. Web camera (Omiky, model: CEL USB 2.0 50.0M PC Camera , catalog number: CEL9002255) with the IR filter removed; Replacing the IR filter with the floppy disk (the black floppy disk contained in the hard shell) to filter the visible light
  5. Behavior room with the temperature and humidity controlled

Software

  1. MATLAB (MathWorks, R2013a)

Procedure

  1. Set the environmental illuminance (provided by the fluorescent lamp) to be 1,000-1,300 Lux, the temperature to be 24 ± 1 °C and the humidity to be 40-60%.
  2. Transfer the flies into the empty vial and anesthetize them on ice for 10 min.
  3. Transfer one fly to each culture dish, and let the flies recover from the anesthesia for at least 10 min.
  4. Record the locomotion of the flies for 2 min with the web camera equipped with a visible light filter (Figures 1A and 1D; Video1). The fly is light up and the background is dark (Figure 1B). The angle of the infrared LEDs should be adjusted before recording to reduce the reflection on the dish.


    Figure 1. Locomotor assay in Drosophila melanogaster. A. The schematic diagram of the locomotor assay; B. A photo of the dish with a fly inside; C. The motion trial of a recorded fly; D. A photo of the setup.

    Video 1. Recording of the locomotor behavior of the flies

    Data analysis

    1. Analyze the photos with MATLAB (MathWorks, R2013a; For MATLAB code for video analysis, see Supplemental file 1) (Figure 1C). Figure out the coordinate of the fly on each photo, and then calculate the moving time, walking length and speed of the fly, and analyze the locomotor trail of the fly.
    2. Individuals with the walk length less than half of the fly body length during recording should be excluded.
    3. The Wilcoxon signed rank test should be applied to evaluate differences between matched samples.

    Acknowledgments

    We would like to thank Jingwu Hou for assistance with experimental setup. This protocol was designed by Q. L. and was implemented in the previously published study (Liu et al., 2016). This study was supported by the ‘Strategic Priority Research Program’ of the CAS (XDB02040004), by grants from the 973 Program (2011CBA00400), as well as by the National Science Foundation of China (91232000, 91132709, 31130027, and 31070956). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

    References

    1. Liu, Q., Yang, X., Tian, J., Gao, Z., Wang, M., Li, Y. and Guo, A. (2016). Gap junction networks in mushroom bodies participate in visual learning and memory in Drosophila. Elife 5.

简介

该协议描述了黑腹果蝇中的简单运动检测。 简单来说,培养皿中的每一只飞行的运动器被网络摄像机记录下来。 可以定量分析单飞的移动时间,步行长度,速度和运动轨迹。
【背景】该协议是在以前发表的研究中实现的(Liu et al。,2016)。 在该研究中,通过简单地提供适当的激发光,将该测定与光遗传系统组合。

关键字:运动, 果蝇, 视频分析, 运动检测, 运动轨迹, 行为

材料和试剂

  1. 3.5厘米培养皿,黑色纸在里面
  2. 要分析的果蝇
  3. 冰用于麻醉

设备

  1. 空瓶用于冷麻醉
  2. 荧光灯(Bannet T5 8 W,中国)
  3. 红外LED(TUOENS,型号:TS-6036A)
  4. 网络摄像机(Omiky,型号:CEL USB 2.0 50.0M PC相机,目录号:CEL9002255),并将IR滤镜取下;用软盘(包含在硬壳中的黑色软盘)更换红外滤光片以过滤可见光
  5. 行为室温湿度控制

软件

  1. MATLAB(MathWorks,R2013a)

程序

  1. 将环境照度(由荧光灯提供)设置为1,000-1,300Lux,温度为24±1℃,湿度为40-60%。
  2. 将苍蝇转移到空的小瓶中,并在冰上麻醉10分钟
  3. 将一只蝇转移到每个培养皿中,让苍蝇从麻醉中恢复至少10分钟。
  4. 用装有可见光滤光片的网络摄像机(图1A和1D; Video1)记录苍蝇的运动2分钟。飞行灯亮起,背景暗暗(图1B)。在记录前应调整红外LED的角度,以减少菜肴上的反射

    图1.黑腹果蝇的运动检测。A.运动检测的示意图;一张内有苍蝇的菜的照片; C.记录飞行的动议审判; D.设置的照片。

    数据分析

    1. 使用MATLAB分析照片(MathWorks,R2013a;对于视频分析的MATLAB代码,请参阅补充文件1 )(图1C)。找出每张照片上飞行的坐标,然后计算飞行的移动时间,步行长度和速度,并分析飞行的运动轨迹。
    2. 记录中步行长度小于飞体长度一半的个人应被排除。
    3. 应用Wilcoxon签名秩检验来评估匹配样本之间的差异。

    致谢

    感谢Jingwu Hou在实验设置方面的帮助。该协议是由Q.L.设计的,并在之前发表的研究中实施(Liu等人,2016)。本研究得到了中国科学院"战略优先研究计划"(XDB02040004),973计划(2011CBA00400)以及中国国家科学基金(91232000,91132709,31130027和31070956)的资助。资助者在研究设计,数据收集和分析,决定出版或准备稿件方面没有任何作用。

    参考

    1. Liu,Q.,Yang,X.,Tian,J.,Gao,Z.,Wang,M.,Li,Y. and Guo,A.(2016)。  蘑菇体中的间隙连接网络参与 D 中的视觉学习和记忆rosophila Elife 5
  • English
  • 中文翻译
免责声明 × 为了向广大用户提供经翻译的内容,www.bio-protocol.org 采用人工翻译与计算机翻译结合的技术翻译了本文章。基于计算机的翻译质量再高,也不及 100% 的人工翻译的质量。为此,我们始终建议用户参考原始英文版本。 Bio-protocol., LLC对翻译版本的准确性不承担任何责任。
Copyright Liu et al. This article is distributed under the terms of the Creative Commons Attribution License (CC BY 4.0).
引用: Readers should cite both the Bio-protocol article and the original research article where this protocol was used:
  1. Liu, Q., Tian, J., Yang, X., Li, Y. and Guo, A. (2017). Locomotor Assay in Drosophila melanogaster. Bio-protocol 7(10): e2283. DOI: 10.21769/BioProtoc.2283.
  2. Liu, Q., Yang, X., Tian, J., Gao, Z., Wang, M., Li, Y. and Guo, A. (2016). Gap junction networks in mushroom bodies participate in visual learning and memory in Drosophila. Elife 5.
提问与回复

(提问前,请先登录)bio-protocol作为媒介平台,会将您的问题转发给作者,并将作者的回复发送至您的邮箱(在bio-protocol注册时所用的邮箱)。为了作者与用户间沟通流畅(作者能准确理解您所遇到的问题并给与正确的建议),我们鼓励用户用图片或者视频的形式来说明遇到的问题。由于本平台用Youtube储存、播放视频,作者需要google 账户来上传视频。

当遇到任务问题时,强烈推荐您提交相关数据(如截屏或视频)。由于Bio-protocol使用Youtube存储、播放视频,如需上传视频,您可能需要一个谷歌账号。