Development of Polymorphic Genic SSR Markers by Transcriptome Sequencing in the Welsh Onion (Allium fistulosum L.)
Development of Polymorphic Genic SSR Markers by Transcriptome Sequencing in the Welsh Onion (Allium fistulosum L.)
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通过转录组测序开发大葱 (Allium fistulosum L.) 的多态性基因 SSR 标记
作者：
Liuyi Yang;Changlong Wen;Hong Zhao;Qianchun Liu;Jingjing Yang;Lecheng Liu;王永勤
转录组分析是探索植物分子标记的一种有效方法，植物基因组序列尚未公布。为了解决大葱分子标记数量有限的问题，本研究利用新一代测序技术（NGS）和RNA-Seq方法，发现了6486个由1-5 bp重复序列组成的基因简单重复序列（SSR）位点。最丰富的基序是单核苷酸（52.33%），其次是三核苷酸（31.96%）和二核苷酸（14.57%）。共设计了2525对引物，在311对引物中有91对引物表现出多态性。总的来说，随机选择了38个SSR标记进一步验证遗传多样性程度，其中22个（57.89%）SSR标记表现出较高的多态性水平。平均多态性信息含量（PIC）值为0.63，等位基因数（Na）为5.27，利用UPGMA聚类分析将22份葱属材料分为3大类，Nei's相似系数在0.37 ~ 0.99之间。这一结果表明，这些基因SSR标记可用于建立更高分辨率的遗传图谱和/或分析葱属植物的系统发育关系在不久的将来。
Transcriptome analysis is an efficient way to explore molecular markers in plant species, for which genome sequences have not been published. To address the limited number of markers published for the Welsh onion, this study found 6486 loci of genic simple sequence repeats (SSR), which consisted of 1–5 bp repeat motifs, based on next-generation sequencing (NGS) technology and the RNA-Seq approach. The most abundant motif was mononucleotide (52.33%), followed by trinucleotide (31.96%), and dinucleotide (14.57%). A total of 2525 primer pairs were successfully designed, and 91 out of 311 tested primers were polymorphisms. Overall, 38 genic SSR markers were randomly selected to further validate the degree of genetic diversity, and 22 genic SSR markers (57.89%) showed high levels of polymorphism. The average polymorphism information content (PIC) value and the number of alleles (Na) were 0.63 and 5.27, respectively, and the unweighted pair-group method with arithmetic average (UPGMA) cluster analysis grouped the 22 Allium accessions into three groups with Nei’s similarity coefficients ranging from 0.37 to 0.99. This result suggested that these genic SSR markers could be used to develop a higher resolution genetic map and/or to analyze the phylogenetic relationships among Allium plants in the near future.
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