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使用斑马鱼对视觉功能进行自动化、高通量的体内分析。

Automated, high-throughput, in vivo analysis of visual function using the zebrafish.

作者信息

Scott C Anthony, Marsden Autumn N, Slusarski Diane C

机构信息

Department of Biology, University of Iowa, Iowa City, Iowa.

Interdisciplinary Graduate Program in Genetics, University of Iowa, Iowa City, Iowa.

出版信息

Dev Dyn. 2016 May;245(5):605-13. doi: 10.1002/dvdy.24398. Epub 2016 Mar 6.

Abstract

BACKGROUND

Modern genomics has enabled the identification of an unprecedented number of genetic variants, which in many cases are extremely rare, associated with blinding disorders. A significant challenge will be determining the pathophysiology of each new variant. The Zebrafish is an excellent model for the study of inherited diseases of the eye. By 5 days post-fertilization (dpf), they have quantifiable behavioral responses to visual stimuli. However, visual behavior assays can take several hours to perform or can only be assessed one fish at a time.

RESULTS

To increase the throughput for vision assays, we used the Viewpoint Zebrabox to automate the visual startle response and created software, Visual Interrogation of Zebrafish Manipulations (VIZN), to automate data analysis. This process allows 96 Zebrafish larvae to be tested and resultant data to be analyzed in less than 35 minutes. We validated this system by disrupting function of a gene necessary for photoreceptor differentiation and observing decreased response to visual stimuli.

CONCLUSIONS

This automated method along with VIZN allows rapid, high-throughput, in vivo testing of Zebrafish's ability to respond to light/dark stimuli. This allows the rapid analysis of novel genes involved in visual function by morpholino, CRISPRS, or small-molecule drug screens. Developmental Dynamics 245:605-613, 2016. © 2016 Wiley Periodicals, Inc.

摘要

背景

现代基因组学已能够识别出数量空前的遗传变异,其中许多变异极为罕见,且与致盲性疾病相关。一项重大挑战将是确定每个新变异的病理生理学机制。斑马鱼是研究眼部遗传性疾病的理想模型。在受精后5天(dpf),它们对视觉刺激具有可量化的行为反应。然而,视觉行为测定可能需要数小时才能完成,或者一次只能评估一条鱼。

结果

为了提高视觉测定的通量,我们使用Viewpoint Zebrabox使视觉惊吓反应自动化,并创建了软件“斑马鱼操作视觉询问”(VIZN)来实现数据分析自动化。这个过程能在不到35分钟内对96条斑马鱼幼虫进行测试并分析所得数据。我们通过破坏光感受器分化所需基因的功能并观察对视觉刺激的反应降低来验证了该系统。

结论

这种自动化方法与VIZN一起,能够对斑马鱼对光/暗刺激的反应能力进行快速、高通量的体内测试。这使得能够通过吗啉代寡核苷酸、CRISPRs或小分子药物筛选快速分析参与视觉功能的新基因。《发育动力学》245:605 - 613,2016年。©2016威利期刊公司。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/da98/4844763/fef84b426803/nihms761762f1.jpg

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