木本油料植物无患子SSR特征分析及SSR-PCR反应体系构建  

SSR characteristics and construction of SSR-PCR reaction system in woody oil plant Sapindus saponaria

作　　者：周宵 蒋丽娟[2] 李昌珠[1] 陈韵竹 李培旺[1] 熊宇晴 张路红 盛克寨 杨艳[1] 陈景震[1] ZHOU Xiao;JIANG Li-juan;LI Chang-zhu;CHEN Yun-zhu;LI Pei-wang;XIONG Yu-qing;ZHANG Lu-hong;SHENG Ke-zhai;YANG Yan;CHEN Jing-zhen(State Key Laboratory of Utilization of Woody Oil Resource,Hunan Academy of Forestry,Changsha 410004,Chi-na;Central South University of Forestry and Technology,College of Life Sciences and Technology,Changsha 410002,China;Hunan Soapberry Agroforestry Development Co.,Ltd.,Changde 415000,China)

机构地区：[1]湖南省林业科学院省部共建木本油料资源利用国家重点实验室,湖南长沙410004 [2]中南林业科技大学生命科学与技术学院,湖南长沙410002 [3]湖南无患子农林发展有限公司,湖南常德415000

出　　处：《中国油料作物学报》2024年第1期72-83,共12页Chinese Journal of Oil Crop Sciences

基　　金：重大科技创新平台项目(2022PT1004);湖南省林业科技计划项目(XLK202101-1);省部共建木本油料资源利用国家重点实验室培育项目(2019XK2002);油脂分子构效湖南省重点实验室项目(2018TP1029);林业科技攻关与创新项目(XLKY202205);中南林业科技大学研究生科技创新基金(CX202101017)。

摘　　要：无患子(Sapindus saponaria L.)是集生物柴油、药用、生态等多种用途于一体的经济型木本油料。本研究利用MISA软件挖掘了无患子叶片转录组中的SSR,对搜索到的SSR进行了特征分析,构建并优化了无患子SSR-PCR反应体系。结果表明,无患子叶片转录组共有Unigene条数目为52460条,其中含有SSR的Unigene有7014条,共检索到8654个SSR。经统计,SSR丰富度从二至六核苷酸依次减少,依次有4327、2669、634、208、161个。SSR长度范围在12~250 bp之间,长度小于等于15 bp的有3619个,均为二和三核苷酸;长度大于15 bp的有5035个,以二、三核苷酸居多。所有SSR共有435种不同重复单元。SSR不同类型重复基元频次分布在4~30次之间,主要分布在4~10频次,共有7090个SSR位点,占总频次的89.65%。SSR位点靶基因GO功能注释显示,无患子叶片转录组Unigene可分为3大类别51个小组;KEGG功能注释显示5大类别中,代谢通路占比最高为65.18%。通过对无患子SSR-PCR反应体系构建与优化,无患子SSR-PCR反应体系最佳组合为:循环次数37次、退火温度60℃、引物量2.0μL、DNA模板浓度为40 ng/μL。Sapindus saponaria L.is an economic tree species for biodiesel,medical,ecological and many other uses.In this study,MISA software was used to mine SSRs in transcriptome of S.saponaria.The SSRs characteristics were analyzed.SSR-PCR reaction system were constructed and optimized.Results showed 52460 unigenes in S.saponaria transcriptome.Among which 7014 unigenes contained SSRs,and a total of 8654 SSRs were discovered.SSR richness was decreased from dinucleotide to hexanucleotide.SSR length ranged from 12 to 250 bp.3619 SSRs had the length less than or equal to 15 bp,all of which were dinucleotide and trinucleotide.The length of the SSRs was greater than 15 bp,mainly dinucleotides and trinucleotides.Totally 435 different repeat units were in these SSRs,including 139,115,109,60,12 and 0 types respectively,from hexanucleotide to mononucleotide.The SSR repeat elements frequencies between 4 to 30 times,89.65%of the total SSRs accounted to 7090 locis were distributed from 4 to 10 times.GO functional annotation of target genes at SSR locis showed that the Unigenes could be divided into 3 categories and 51 groups.KEGG function annotation showed that the metabolism pathway accounted for 65.18%in 5 categories.Through the construction and optimization of the reaction system of S.saponaria SSR-PCR,the optimal combination was as follows:37 cycles,annealing temperature 60℃,primer amount 2.0μL and DNA template concentration 40 ng/μL.

关 键 词：木本油料植物 EST-SSR 功能注释 PCR反应体系 生物柴油 

分 类 号：S727.1[农业科学—林木遗传育种] S759.33[农业科学—林学]