基于IRAP标记的沙子空心李遗传多样性评价及指纹图谱构建
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作者:
作者单位:

1.贵州大学生命科学学院/山地植物资源保护与种质创新教育部重点实验室,贵阳 550025;2.贵州省铜仁市林业科学院,铜仁 554300;3.贵州省铜仁市农业科学院,铜仁 554300;4.贵州省农业科学院,贵阳 550006

作者简介:

王贵,E-mail:2051647320@qq.com

通讯作者:

乔光,E-mail:13518504594@163.com

中图分类号:

S662.3

基金项目:

贵州省科技计划项目(黔科合支撑[2020]1Y023);贵州省科技计划项目(黔科合服企[2019]4013);中央引导地方科技发展资金项目(黔科中引地[2021]4010)


Evaluation of genetic diversity and construction of DNA fingerprint of Prunus salicina Lindl. ‘Shazikongxinli’ based on IRAP markers
Author:
Affiliation:

1.The Key Laboratory of Plant Resources Conservation and Germplasm Innovation in Mountain Region(Ministry of Education)/College of Life Sciences, Guizhou University,Guiyang 550025,China;2.Tongren Academy of Forestry,Tongren 554300,China;3.Tongren Academy of Agricultural Sciences,Tongren 554300,China;4.Guizhou Academy of Agricultural Sciences,Guiyang 550006,China

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    摘要:

    通过走访调查,选取92株具有优良性状的沙子空心李种质,根据李反转录转座子的RT序列开发IRAP标记引物,设计L16( 43 ) 的正交试验优化10 μL IRAP-PCR主要影响因素的含量;利用18条IRAP引物对92份沙子空心李进行遗传多样性分析,构建供试种质的指纹图谱。结果显示:最优的10 μL IRAP-PCR反应体系为10-5 mol/L IRAP引物l.3 μL,PCR Mix 5.0 μL、模板DNA(30 ng/μL) l.0 μL,PCR循环40次。18条IRAP引物共扩增出189个位点,多态性位点180个,等位基因数、有效等位基因数、Nei’s基因多样性指数和Shannon指数均值分别为1.930、1.426、0.261及0.405。采用邻接法将92份沙子空心李聚类为4个类群。确定IRAP引物Ty3-2及Ty3-6为核心引物,可高效构建92份供试种质的指纹图谱。

    Abstract:

    Ninty-two accessions of ‘Shazikongxinli’ germplasm with elite traits were selected by visiting and surveying to evaluate the genetic diversity of Prunus salicina Lindl. ‘Shazikongxinli’ IRAP primers were developed based on the conserved regions of RT sequences of plum reverse transcription transposons Ty1-copia and Ty3-gypsy. L16(43) orthogonal experiments were designed to optimize the main factors affecting content of IRAP-PCR with 10 μL volume. 18 IRAP primers were used to analyze the genetic diversity and construct the fingerprint of the germplasm tested. The results showed that the optimized 10 μL IRAP-PCR reaction system was 1.3 μL of 5-10 mol/L IRAP primers, 5.0 μL of PCR mixture, 1.0 μL of template DNA(30 ng/μL), and the number of PCR cycles was 40 times. A total of 189 loci were amplified from 18 IRAP primers with good repeatability and high polymorphism, including 180 polymorphic loci.The average value of observed number of alleles(Na), effective number of alleles(Ne), Nei’s gene diversity(He) and Shannon’s information index(I) for all primers was 1.930,1.426,0.261 and 0.405, respectively. The 92 accessions tested were clustered into 4 groups using the neighbor-joining method. The IRAP primers Ty3-2 and Ty3-6 were identified as core primers, which can be used to efficiently construct the fingerprints of 92 accessions tested.

    表 4 IRAP引物扩增结果统计Table 4 Statistics of IRAP primer amplification results
    图1 10 μL IRAP-PCR体系优化结果Fig.1 Optimization results of 10 μL IRAP-PCR system
    图2 6条IRAP引物筛选结果Fig.2 Screening results of 6 IRAP primers
    图3 引物Ty3-2及Ty3-6 PCR扩增结果Fig.3 PCR amplification results of Ty3-2 and Ty3-6
    图4 92份沙子空心李聚类结果Fig.4 Dendrogram of 92 Shazikongxinli germplasms based on IRAP bands
    图5 沙子空心李种质N1及L1分子身份证Fig.5 Prunus salicina Lindl. Shazikongxinli N1 and L1 molecular IDs
    表 5 IRAP引物鉴别供试种质情况Table 5 The number of germplasms that can be identified by IRAP primers
    表 2 L16( 43 ) 正交试验表Table 2 Table of L16(43) orthogonal experiments
    表 1 92份沙子空心李种质材料Table 1 Ninety-two germplasm resources of Shazikongxinli for IRAP analysis
    表 3 18条IRAP引物信息Table 3 Information of 18 IRAP primers
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王贵,李蕊蕊,吴茂宏,任菲宏,王丽丽,乔光.基于IRAP标记的沙子空心李遗传多样性评价及指纹图谱构建[J].华中农业大学学报,2023,42(1):1-11

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  • 收稿日期:2022-08-30
  • 在线发布日期: 2023-02-22
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