基于转录组数据高通量发掘沙葱萤叶 甲微卫星引物
High-throughput discovery of microsatellite markers in Galeruca daurica (Coleoptera: Chrysomelidae) from a transcriptome database
张鹏飞1** 周晓榕1 庞保平1*** 谭 瑶1 常 静1 高利军2
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DOI：10.7679/j.issn.2095-1353.2016.130
作者单位：1. 内蒙古农业大学草原昆虫研究中心，呼和浩特 010019；2. 锡林浩特市草原工作站，锡林浩特 026000
中文关键词：沙葱萤叶甲，微卫星，高通量测序，转录组
英文关键词：Galeruca daurica, microsatellite, high-throughput sequencing, transcriptome
中文摘要：
【目的】 沙葱萤叶甲Galeruca daurica（Joannis）是近年来在内蒙古草原上猖獗发生为害的一种新害虫。本研究从其高通量转录组数据中查找微卫星序列，并进行微卫星位点的信息分析和微卫星引物的发掘，将为进一步研究沙葱萤叶甲遗传多样性及遗传分化奠定必要的基础。【方法】 应用软件MISA对沙葱萤叶甲转录组中72 352条unigenes的数据进行搜索。【结果】 共找到3 880个微卫星位点，分布于3 277条unigenes中，其主要重复类型为单核苷酸重复（80.85%），其次为三核苷酸重复（11.08%），再次为二核苷酸重复（7.37%）。单核苷酸重复中主要是A/T基序，占总量的77.76%。从1 814条unigenes 中成功设计出2 160对引物，从中随机选取10对引物对沙葱萤叶甲DNA进行扩增，结果全部扩增出目的DNA片段。【结论】 利用沙葱萤叶甲转录组数据开发微卫星引物是可行的，本研究开发的微卫星引物为研究沙葱萤叶甲种群遗传学和功能基因组学奠定了必要的基础。
英文摘要：
[Objectives] Galeruca daurica (Joannis) is a new pest which has had a serious impact on Inner Mongolian grasslands in recent years. We investigated microsatellite sequences from its high-throughput transcriptome database, analyzed microsatellite loci information, and developed microsatellite primers in order to make an essential foundation for further study of the genetic diversity and differentiation of G. daurica populations. [Methods] We used the software MISA to search for microsatellite sequences among 72 352 unigenes of the G. daurica transcriptome database. [Results] 3 880 microsatellite loci distributed in 3 277 unigenes were identified. The main repeat types were mono-nucleotide repeats (80.85%), followed by tri-nucleotide repeats (11.08%) and finally bi-nucleotide repeats (7.37%). The A/T motif was the most abundant (77.75%) in mono-nucleotide repeats. Based on identified microsatellite sequences, 2 160 pairs of primers were designed from 1 814 unigenes, 10 pairs of which were randomly selected and used to amplify the G. daurica DNA. All 10 pairs of primers could amplify the target DNA fragments. [Conclusion] It is feasible to develop microsatellite primers from the G. daurica transcriptome. The primers developed here provide an essential foundation for studying the population genetics and functional genomics of G. daurica.