生物学杂志

生物学杂志 ›› 2019, Vol. 36 ›› Issue (5): 11-.doi: 10.3969/j.issn.2095-1736.2019.05.011

• 研究报告 • 上一篇    下一篇

斑马鱼hoxb7a基因在hox基因簇中的作用研究

  

  1. １．上海海洋大学科技部海洋生物科学国际联合研究中心， 上海 201306；
    ２．上海海洋大学水产种质资源发掘与利用教育部重点实验室， 上海 201306； 
    ３．上海海洋大学水产科学国家级实验教学示范中心， 上海 201306
  • 出版日期:2019-10-18 发布日期:2019-10-11
  • 通讯作者: 祖尧，副教授，硕士生导师，研究方向为遗传与发育生物学，E-mail: yzu@shou.edu.cn
  • 作者简介:孟祥丽，硕士研究生，研究方向为遗传与发育生物学，E-mail: m150110208@st.shou.edu.cn
  • 基金资助:
    国家自然基金青年基金（31501166）；上海市人力资源和社会保障局上海市人才发展资金（201568）；上海市教委晨光计划（14CG49）

The function study of hoxb7a in hox clusters in zebrafish

  1. 1. International Research Center for Marine Biosciences, Ministry of Science and Technology, Shanghai Ocean University, Shanghai 201306; 2. Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources, Ministry of Education, Shanghai Ocean University, Shanghai 201306； 3. National Demonstration Center for Experimental Fisheries Science Education, Shanghai Ocean University, Shanghai 201306, China
  • Online:2019-10-18 Published:2019-10-11

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摘要： 斑马鱼hoxb7a基因是hox 基因家族在进化过程中第7旁系同源组中仅存的基因。为了研究hoxb7a基因在整个hox基因簇中的作用，利用CRISPR/Cas9技术对斑马鱼hoxb7a基因进行编辑，T7E1结果显示F0敲除效率平均为43%，并在F2成功获得缺失8 个碱基(Δ8 bp)和缺失23 个碱基(Δ23 bp)的 hoxb7a纯合突变体。进而利用实时荧光定量PCR技术检测48hpf野生型和hoxb7a突变体中旁系同源hox基因的表达量变化，QPCR结果显示，hoxb7a缺失后hox基因簇中相邻的hox基因表达量显著性升高（P<0.001），而基因组上距离hoxb7a较远的hox基因表达量未出现明显变化（P>0.05）。研究表明hoxb7a基因在整个hox基因簇中发挥重要调控作用，通过阐述hoxb7a的作用加深了对hox基因簇加倍后基因功能分化的认识。同时斑马鱼hoxb7a突变体的成功构建，为其基因功能机制的研究奠定了坚实基础。

关键词: font-size:medium, ">斑马鱼；CRISPR/Cas9系统；hoxb7a基因；hox基因簇；旁系同源基因

Abstract: The zebrafish hoxb7a gene is the only gene in the seventh paralogous group of hox genes in the evolution of the third genome duplication. In order to study the function of hoxb7a gene in the whole hox gene clusters, we knocked out the zebrafish hoxb7a by using CRISPR/Cas9 technology. The hoxb7a mutagenesis efficiency was about 43% which was evaluated by T7E1 assay in F0. And we successfully obtained two mutant fish lines of hoxb7a, which were 8 bp deletion and 23 bp deletion. Furthermore, we used QPCR technology to compare the related hox genes expression level changing of wide type zebrafish embryos with that of hoxb7a-/-at 48hpf (hour post fertilization). And the QPCR results showed that the hox genes closed to hoxb7a in the genome were significantly up-regulated (P<0.001), however, the hox genes far away from hoxb7a in the genome had no obvious difference(P>0.05)in hoxb7a mutants comparing with the wide type. This study demonstrated that hoxb7a plays a crucial role in hox clusters, which deepens the understanding of the functional differentiation of duplicate hox genes. At the same time, we successfully established the hoxb7a mutant, which provides a foundation for future mechanism studies on the hoxb7a gene function.

Key words: zebrafish, CRISPR/Cas9 system, hoxb7a gene, hox clusters, paralogous gene

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