Development, characterization, and annotation of potential simple sequence repeats by transcriptome sequencing in pears (Pyrus pyrifolia Nakai).
Development, characterization, and annotation of potential simple sequence repeats by transcriptome sequencing in pears (Pyrus pyrifolia Nakai).
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DOI：
10.4238/gmr.15038683
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发表时间：
2016-09
期刊：
影响因子：
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通讯作者：
H. Zhou;Bin Cai;Z. Q. Lu;Zhihong Gao;Yushan Qiao
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文献类型：
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作者：
H. Zhou;Bin Cai;Z. Q. Lu;Zhihong Gao;Yushan Qiao
简单重复序列（SSR）是最有效的分子标记之一，可用于DNA指纹图谱、品种鉴定、遗传作图和标记辅助选择。利用梨（Pyrus pyrifolia Nakai）的75，764个单基因（55，676，271 bp）进行深度转录组测序，在9154个单基因中鉴定出10，622个新的SSR，占单基因总数的14.02%。这些SSR的平均长度为16 bp，平均分布为5.24 kb。二核苷酸重复基序是主要的类型，频率为55.87%，其次是三核苷酸重复基序（24.45%）。梨转录组中存在159种重复基序。AG/CT是最常见的基序，占49.64%。使用Nr（NCBI非冗余蛋白质数据库）、Nt（NCBI非冗余核苷酸数据库）和Swiss-Prot数据库对所有9154个含有SSR的单基因进行功能注释，并通过Gene Ontology和Clusters of Orthopathic Groups进一步分类。此外，共设计了4300对引物。随机选取40个引物进行PCR扩增和聚丙烯酰胺凝胶电泳（PAGE）分析。在40对引物中，有31对引物经PAGE电泳分离成功。这些发现也证实了使用下一代测序技术挖掘SSR是一种快速，有效和可靠的方法。
Simple sequence repeats (SSRs), one of the most powerful molecular markers, can be used for DNA fingerprinting, variety identification, genetic mapping, and marker-assisted selection. Using the pear's (Pyrus pyrifolia Nakai) 75,764 unigenes (55,676,271 bp) obtained by deep transcriptome sequencing, a total of 10,622 novel SSRs were identified in 9154 unigenes, accounting for 14.02% of all unigenes. The average length and distribution of these SSRs was about 16 bp and 5.24 kb, respectively. Dinucleotide repeat motifs were the main type, with a frequency of 55.87%, followed by trinucleotides (24.45%). There were 159 kinds of repeat motifs existing in the pear transcriptome. AG/CT was the most frequent motif, accounting for 49.64%. All 9154 SSR-containing unigenes were functionally annotated using Nr (NCBI non-redundant protein database), Nt (NCBI non-redundant nucleotide database), and the Swiss-Prot database, and were classified further by Gene Ontology and Clusters of Orthologous Groups. In addition, a total of 4300 primer pairs were designed from all SSR loci obtained. Of these, 40 primers were randomly selected for PCR amplification and polyacrylamide gel (PAGE) analysis. Among the 40 primer pairs, 31 were successfully separated via PAGE. These findings also confirm that mining SSRs using next-generating sequencing technologies is a fast, effective, and reliable approach.