基于基因组数据库的杂草稻叶绿体分子标记开发  

Development of molecular markers in weedy rice chloroplast based on genomic database

作　　者：朱国忠[1] 戴伟民[1] 陈晓锋[1] 张晶旭[1] 强胜[1] 

机构地区：[1]南京农业大学杂草研究室/江苏省杂草防治技术工程技术研究中心,江苏南京210095

出　　处：《南京农业大学学报》2015年第2期240-247,共8页Journal of Nanjing Agricultural University

基　　金：国家转基因生物新品种培育重大专项(2014ZX08011);教育部高等学校博士点基金优先发展领域课题(20130097130006)

摘　　要：[目的]杂草稻的起源与演化是一个未澄清的热点问题,分子标记技术是最重要的研究手段之一。由于已经应用的分子标记数量有限,与栽培水稻缺乏特异性,因此探索新的分子标记仍然十分必要。[方法]使用网络基因组数据库中13份栽培水稻和野生稻叶绿体全基因组序列进行比对和分析,针对其中的高频变异区域设计标记引物,进而对60份杂草稻样品的标记区域测序并进行多态性验证。[结果]13份样品叶绿体全基因组中存在1020个单核苷酸多态性(SNP)位点,111个插入/缺失(InDel)位点,表现出丰富的遗传变异;选定其中3个高频变异区域,分别针对trnG-trnfM基因间区、psbM-trnC基因间区和trnT-trnL基因间区设计了3对引物,对60份杂草稻材料进行了检测,发现3个片段总长1966bp,共存在7个SNP位点,6个InDel位点,总变异序列长度为34bp,占序列总长的1.73%。系统发育树结果显示,60份杂草稻被划分为4个类群。另外,将网络数据库中的13份栽培水稻和野生稻数据进行分析,发现不同栽培稻和野生稻的籼粳类型分别被聚到杂草稻的相应类群中。[结论]所开发的3对叶绿体分子标记可以对上述杂草稻进行良好的分类,因此可以作为研究杂草稻遗传多样性及演化的分子标记。[Objectives]The origin and evolution of weedy rice still is a disputed but hot issue.The molecular markers applied in a study of weedy rice are limited and not specific to weedy rice discriminating from cultivated rice.Hence,it is necessary to explore new molecular markers in this field.[Methods]The downloaded chloroplast genome sequences of 13 cultivated rice and wild rice from genomic database were aligned and analyzed.We designed primers as markers based on the sequence of the high frequency variation regions.Then the labeling areas in 60 accessions of weedy rice were sequenced and verified for the polymorphism.[Results]The results showed that there were 1 020 SNPs and 111 In Dels in the chloroplastic genome sequences of 13 cultivated rice and wild rice,which had rich genetic variation.Three primers were designed as markers in the selected 3 high frequency variation regions which were located in trn G-trnf M inter spacer,psb M-trn C inter spacer and trn T-trn L inter spacer.The detection of 60 weedy rice accessions using the developed three markers demonstrated that there were 7 SNPs and 6 In Dels in these three regions with a total sequence of 1 966 bp among the weedy rice.The total length of the variation sequence contained 34 bp with 1.73% variation of the whole sequence length.The phylogenetic tree constructed by tag sequence divided the 60 weedy rice accessions into four groups.Additionally,taking 13 cultivated rice and wild rice sequences from the network database,it was found that japonica and indica rice and cultivated rice were always clustered into the corresponding weedy rice classes.[Conclusions]The three markers developed in this study could well classify weedy rice accessions and showed a potential application for weedy rice genetic diversity and evolution research.

关 键 词：杂草稻 基因组数据库 叶绿体基因 分子标记 

分 类 号：Q943.2[生物学—植物学] S451.1[农业科学—植物保护]