斑马鱼小头震颤突变体 mise 的鉴定与基因定位
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国家卫生健康委科学技术研究所实验动物中心,北京 100081

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R-33

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Identification and gene localization of a microcephaly and microcephaly and suspected epilepsy mutant mise in zebrafish
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Experimental Animal Center of National Research Institute for Family Planning, Beijing 100081, China

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    摘要:

    目的 以自发突变导致的小头、震颤(microcephaly and seizure,mise)斑马鱼为模型,研究其遗传方式和突变基因定位。 方法 通过比较正常和突变体斑马鱼的头宽和眼球面积,鉴定突变体的小头小眼性状;通过构建家系确定突变遗传方式。对极端表型群体进行基因组重测序,随后利用分离群体分组分析( bulked segregation analysis,BSA)和竞争性等位基因特异性 PCR(kompetitive allele specific PCR,KASP),一代测序等方法获得突变基因定位。 结果 与野生型相比,突变体 mise 受精后 3 d(3 days post fertilization,3 dpf)表现为小头小眼震颤表型。经基因定位及验证明确与表型相关的突变基因为 terfa。 结论 本研究通过全基因组重测序结合连锁分析的方法快速定位突变基因,可为斑马鱼突变体的基因定位提供参考,为神经系统发育异常的相关基因功能分析奠定基础。

    Abstract:

    Objective Taking the congenital microcephaly and suspected epilepsy (mise)zebrafish mutant caused by spontaneous as a model, to study its inheritance pattern and phenotype-related mutation gene location. Methods By comparing the head width and eye area of normal and mutant zebrafish, the microcephaly and ommatidia traits of mutants were identified; the inheritance pattern of the mutation was determined by constructing a family. Genome resequencing combined with bulked segregation analysis (BSA) and kompetitive allele specific PCR ( kompetitive allele specific PCR, KASP)and Sanger sequencing were used to obtain the location of mutant genes. Results Compared with the wild type, the head and eye were significantly reduced at 3dpf, accompanied by convulsions and tremors. The mutant gene associated with phenotype was identified as terfa by gene mapping and verification. Conclusions This study uses whole-genome resequencing combined with linkage analysis to quickly locate mutant genes. It laid the foundation for the locational cloning of zebrafish mutants and the functional analysis of genes related to abnormal development of the nervous system.

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何嘉玲,王天奇,暴 国,张长勇,李 楠,孙德明.斑马鱼小头震颤突变体 mise 的鉴定与基因定位[J].中国比较医学杂志,2022,32(8):48~56.

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  • 收稿日期:2021-12-15
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  • 在线发布日期: 2022-12-29
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