| 基于转录组测序的圆口铜鱼微卫星标记筛选 |
| Development of Microsatellite Markers for Coreius guichenoti Based on Transcriptome Sequencing |
| 投稿时间：2019-11-07 修订日期：2021-07-22 |
| DOI：10.15928/j.1674-3075.201911070275 |
| 中文关键词:圆口铜鱼 转录组测序 微卫星标记 |
| 英文关键词:Coreius guichenoti transcriptome sequencing microsatellite markers |
| 基金项目:国家重点研发计划项目（No.2019YFD0900603）；现代农业产业技术体系专项资金 (CARS-46)；院级基本科研业务经费(2020TD57) |
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| 中文摘要: |
| 分析圆口铜鱼(Coreius guichenoti)的遗传多样性，可为其人工繁殖家系鉴别、分子标记辅助育种及长江上游珍稀特有鱼类的种质资源保护提供理论依据和技术支撑。以室内循环水养殖的圆口铜鱼为研究对象，提取其肝脏RNA，通过Illumina HiSeq? 2000 高通量测序平台进行转录组测序，利用MISA软件对测序获得的unigene进行微卫星（SSR）位点挖掘和特征分析，设计SSR位点引物并进行验证。结果显示，圆口铜鱼转录组测序共获得80 688条 unigene，通过查找获得34 155个SSR位点，分布在25 947条序列上，发生率为32.2%。圆口铜鱼SSR中主要重复单元类型为单碱基和二碱基，其中A/T和AC/GT为优势单碱基和二碱基重复单元，占总SSR数量的57.2%和19.7%。SSR重复数在5~60次，其中有40.3%的SSR序列集中在11~15次；SSR序列长度变化显著，单碱基至六碱基重复类型的总体长度在11~101 bp，其中12~35 bp序列长度占78.8%。随机选取50个SSR位点合成引物对9尾圆口铜鱼样本进行PCR 扩增验证，可稳定扩增出目的条带的有40对引物，其中16对具有多态性，证明了通过转录组测序开发圆口铜鱼SSR标记的可行性。 |
| 英文摘要: |
| Largemouth bronze gudgeon (Coreius guichenoti) belongs to Coreius, Gobioninae and Cypriniformes. As an indicator species of rare and endemic fish, C. guichenoti is primarily distributed in the upper reaches of the Yangtze River. In this study, RNA was extracted from the liver of C. guichenoti cultured in an indoor recirculating aquaculture system, and transcriptome sequencing was conducted using the Illumina HiSeq? 2000 sequencing platform. The simple sequence repeats (SSR) loci were extracted and analyzed by MISA and then SSR loci primers were designed, tested and verified. A total of 80 688 unigenes were obtained from the transcriptome sequencing of C. guichenoti and 34 155 SSR loci were obtained, distributed in 25 947 unigenes at a frequency of 32.2%. Mononucleotides and dinucleotides were the primary repeat units and A/T and AC/GT were the dominant repeat units with proportions of 57.2% and 19.7% of the total SSR. The number of SSR repeats ranged from 5-60, with 11-15 repeats accounting for 40.3%. The length of SSR sequence changed significantly and ranged from 11-101 bp, with lengths of 12-35 bp accounting for 78.8%. Fifty SSR loci primers were randomly selected and validated in nine C. guichenoti samples. Most of the primers (40) amplified well and yielded clear and readable products, and 16 primers were polymorphic, proving that SSR markers of C. guichenoti could be developed by transcriptome sequencing. The results provide a theoretical basis and technical guidance for analyzing genetic diversity and molecular marker-assisted breeding of C. guichenoti. |
| 李学梅,吴兴兵,杨德国,朱永久.2021.基于转录组测序的圆口铜鱼微卫星标记筛选[J].水生态学杂志,42(4):97-103. |
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