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    洪文娟, 郝兆祥, 刘康佳, 罗华, 毕润霞, 苑兆和, 宗世祥, 王君. 基于石榴全基因组序列的SSR标记开发及鉴定[J]. 北京林业大学学报, 2019, 41(8): 38-47. DOI: 10.13332/j.1000-1522.20190167
    引用本文: 洪文娟, 郝兆祥, 刘康佳, 罗华, 毕润霞, 苑兆和, 宗世祥, 王君. 基于石榴全基因组序列的SSR标记开发及鉴定[J]. 北京林业大学学报, 2019, 41(8): 38-47. DOI: 10.13332/j.1000-1522.20190167
    Hong Wenjuan, Hao Zhaoxiang, Liu Kangjia, Luo Hua, Bi Runxia, Yuan Zhaohe, Zong Shixiang, Wang Jun. Development and identification of SSR molecular markers based on whole genomic sequences of Punica granatum[J]. Journal of Beijing Forestry University, 2019, 41(8): 38-47. DOI: 10.13332/j.1000-1522.20190167
    Citation: Hong Wenjuan, Hao Zhaoxiang, Liu Kangjia, Luo Hua, Bi Runxia, Yuan Zhaohe, Zong Shixiang, Wang Jun. Development and identification of SSR molecular markers based on whole genomic sequences of Punica granatum[J]. Journal of Beijing Forestry University, 2019, 41(8): 38-47. DOI: 10.13332/j.1000-1522.20190167

    基于石榴全基因组序列的SSR标记开发及鉴定

    Development and identification of SSR molecular markers based on whole genomic sequences of Punica granatum

    • 摘要:
      目的利用cDNA文库筛选的石榴SSR多态性标记数量有限,为进一步推动石榴遗传多样性分析、遗传图谱构建、品种鉴定等研究,有必要系统开发高效稳定的分子标记位点。
      方法本研究根据已发表的石榴基因组测序数据利用MISA软件对1 ~ 6核苷酸重复的SSR位点进行了查找,分析了不同类型SSR位点的序列特征,进而设计引物并检测了引物的有效性和多态性。
      结果(1)石榴基因组中共检测到146 445个SSR位点,其中以单核苷酸重复型SSR最多(占51.95%),六核苷酸重复型最少(仅占0.38%);SSR序列以A/T碱基占主导,具有偏向性。(2)石榴基因组SSR序列长度变化范围为10 ~ 252 bp,平均长度15.48 bp。不同长度重复单元类型的SSR序列长度存在丰富变异,呈现随着重复次数增多,SSR序列丰度减少的趋势。(3)根据不同类型SSR位点设计并合成引物140对,其中119对在12份石榴种质中可扩增出有效条带,41对可产生多态性条带,PIC值在0.007 ~ 0.566之间;从中筛选出多态性较高、稳定性好的引物15对,共检测到等位基因44个,平均每个SSR位点检测到2.933 3个等位基因。
      结论利用石榴全基因组序列可实现SSR标记的大规模开发,并可鉴定出大量适用于石榴遗传多样性分析、遗传图谱构建、品种鉴定等研究的SSR引物。相关研究为石榴的遗传育种研究提供了丰富的SSR序列信息和标记资源。

       

      Abstract:
      Objective The number of polymorphic SSR markers developed through cDNA library is limited. In order to promote studies on genetic diversity, genetic mapping and variety identification, it is necessary to develop molecular marker loci.
      Method In this study, SSRs with 1−6 nucleotide repeats were searched from the published whole genome sequences of Punica granatum using MISA and the characterization of SSR sequences and their polymorphisms were analyzed.
      Result (1) A total of 146 445 SSR loci were detected in whole genome sequences of Punica granatum. Then content of mono-nucleotide repeat SSRs was the highest, accounting for 51.95%; in contrast, the content of hexa-nucleotide repeat SSRs was the lowest, only 0.38%. In all SSR sequences, most of bases were A/T, showing a bias in SSR sequences. (2) The range of SSR length ranged from 10 to 252 bp, with an average of 15.48 bp. The SSR length among different types of SSRs varied a lot, and the abundance of SSR sequences tended to decrease with the increase of repeat number. (3) In designed 140 pairs of primers including different types of SSRs, valid fragments could be produced with 119 pairs in 12 germplasms of Punica granatum and 41 pairs could produce polymorphic fragments with 0.007−0.566 polymorphic information content (PIC). We further screened 15 pairs of primers with polymorphic and stable fragments, which detected 44 alleles in these germplasms of Punica granatum, with average of 2.933 3 alleles per SSR locus.
      Conclusion SSR markers can be developed in large-scale based on whole genome sequences of Punica granatum and a large number of SSR primers also can be identified for genetic diversity analysis, genetic mapping and variety identification. This study provides lots of SSR information and marker resources for researching genetics and breeding of Punica granatum.

       

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