| A Modified Enrichment Method to Construct Microsatellite Library from Plateau Pika Genome （Ochotona curzoniae） |
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| 作者单位： | [1]CAS Key Laboratory of Genome Sciences and lnformation, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100029, China [2]Northwest Plateau Institute of Biology, Chinese Academy of Sciences, Xining 810001, China [3]Graduate University of Chinese Academy of Sciences, BeiI＇ing 100049, China. |
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| 摘 要： |
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| 关 键 词： | 基因组文库 微卫星重复 高原鼠兔 富集方法 DNA片段 传统方法 阳性克隆 中国仓鼠 |
A Modified Enrichment Method to Construct Microsatellite Library from Plateau Pika Genome(Ochotona curzoniae) |
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| Authors: | Jianing Geng Kexin Li Yanming Zhang Songnian Hu |
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| Institution: | 1.CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100029, China;2.Northwest Plateau Institute of Biology, Chinese Academy of Sciences, Xining 810001, China;3.Graduate University of Chinese Academy of Sciences, Beijing 100049, China |
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| Abstract: | A microsatellite-enriched library of plateau pika(Ochotona curzoniae)was constructed according to the strong affinity between biotin and streptavidin.Firstly,genomic DNA was fragmented by ultrasonication,which is a major improvement over traditional methods.Linker-ligated DNA fragments were hybridized with biotinylated microsatellite probes,and then were subjected to streptavidin-coated magnetic beads.PCR amplification was performed to obtain double-stranded DNA fragments containing microsatellites.Ligation and transformation were carried out by using the pGEM-T Vector System Ⅰ and Escherichia coli DH10B competent cells.Sequencing results showed that 80.2% of clones contained microsatellite repeat motif.Several modifications make this protocol time-efficient and technically easier than the traditional ones; particularly,composition and relative abundance of microsatellite repeats in plateau pika genome were truly represented through the optimized PCR conditions.This method has also been successfully applied to construct microsatellite-enriched genomic libraries of Chinese hamster(Cricetulus griseus)and small abaloneHaliotis diversicolor(Reeve)]with high rates of positive clones,demonstrating its feasibility and stability. |
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| Keywords: | microsatellite enriched library magnetic bead ultrasonication |
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