鲫鱼基因组DNA的扩增及在SSR分析中的验证  

Increase of Genomic DNA with Verification by SSR Analysis in Carp Carassius auratus

作　　者：武祥伟[1,3] 罗荣 荣华 王晓雯[1,3] 邓君明 孔令富[1,3] 毕保良 WU Xiangwei;LUO Rong;RONG Hua;WANG Xiaowen;DENG Junming;KONG Lingfu;BI Baoliang(Faculty of Animal Science and Technology,Yunnan Agricultural University,Kunming 650201,China;Aquaculture Technology Extension Station of Zhenkang,Zhenkang 677704,China;Key Laboratory of Plateau Fishery Resources Protection and Sustainable Utilization of Yunnan Province,Kunming 650201,China)

机构地区：[1]云南农业大学动物科学技术学院,云南昆明650201 [2]云南省临沧市镇康县水产技术推广站,云南镇康677704 [3]云南省高原渔业资源保护与可持续利用重点实验室,云南昆明650201

出　　处：《云南农业大学学报（自然科学版）》2019年第6期988-993,共6页Journal of Yunnan Agricultural University:Natural Science

基　　金：云南省科技厅应用基础研究项目（2014FD019）

摘　　要：【目的】优化鲫鱼全基因组DNA扩增方法。【方法】使用内切酶酶切初孵仔鱼的基因组DNA,酶切片段连接寡核苷酸接头,根据酶切位点序列与接头序列设计引物,采用PCR法扩增酶切片段,增加基因组DNA的数量。【结果】使用Taq I内切酶酶切鲫鱼基因组DNA,酶切片段连接寡核苷酸接头后PCR扩增,获得扩增产物集中于250~1 500 bp。以扩增的滇池高背鲫鱼基因组DNA为模板,PCR扩增滇池高背鲫鱼的微卫星分子标记(SSR)位点,获得预期的扩增片段,电泳图谱与对照组(未扩增的基因组DNA为PCR模板)无差异。【结论】本研究优化了鲫鱼基因组DNA的扩增方法,并可用于SSR分析中,为鱼类大规模遗传分析提供了技术支撑。[Purpose]The purpose of the present study is the development and optimization of a method for genomic DNA increase in carp Carassius auratus.[Method]The first step for genomic DNA increase was digestion for genomic DNA by endonuclease. Subsequently, the DNA fragments were linked with specific oligonucleotide adaptors. The DNA fragments were amplified by PCR using the primers that were designed from the sequences of endonuclease recognition sites and adaptors.Finally, the genomic DNA increased.[Result]The genomic DNA was digested by Taq I endonuclease. Those fragments were linked with specific oligonucleotide adapotors and were amplified by PCR method. The length of PCR product ranged from 250 bp to 1 500 bp for the major DNA fragments. The genomic DNA of carp C. auratus increased by the developed method and was used for microsatellite loci(SSR) amplification. The expected PCR product was obtained for the SSR loci. Compared to the control(genomic DNA was used for PCR), their electrophoresis patterns were the same.[Conclusion]The present study developed a method for genomic DNA increase by PCR in carp C. auratus, which can be applied for SSR analysis and provides the technology support for the massive genetic analysis in fish species.

关 键 词：滇池高背鲫 基因组DNA PCR 微卫星 内切酶 

分 类 号：S917.4[农业科学—水产科学]