利用iPBS技术克隆牡丹反转录转座子LTR序列  

iPBS-PCR Used for Cloning and Analysis of Long Terminal Repeat Transposons in Tree Peony(Paeonia)

作　　者：张曦[1] 侯小改[1,2] 郭大龙[3,2] 宋程威[1] 段亚宾 

机构地区：[1]河南科技大学农学院,洛阳471003 [2]河南省高校牡丹工程技术中心,洛阳471003 [3]河南科技大学林学院,洛阳471003

出　　处：《植物学报》2014年第3期322-330,共9页Chinese Bulletin of Botany

基　　金：国家自然科学基金(No.31070620);河南省高校科技创新人才支持计划(No.13HASTIT004);河南省重点科技攻关项目(No.132102110029)

摘　　要：利用iPBS方法从西北牡丹(Paeonia suffruticosa)品种红绣球和中原牡丹品种洛阳红中扩增出相应片段,经回收、克隆及测序,获得了12条来自牡丹LTR类反转录转座子的LTR序列,并用相关生物信息学软件对序列进行分析。结果表明,这些核苷酸序列表现出较高的异质性,主要表现为缺失突变,序列长度变化范围为313–894 bp,同源性从31.1%–65.8%不等。将其氨基酸序列与已登录的不同植物LTR类反转录转座子LTR氨基酸序列进行聚类分析,结果显示与某些植物相应序列具有较高的同源性,表明可能存在LTR类反转录转座子的横向传递关系。根据克隆出的LTR序列设计SSAP引物,对牡丹29个品种进行了SSAP分子标记分析,结果显示具丰富的多态性。实验验证了用iPBS技术分离牡丹LTR序列的适用性,并为牡丹种质资源评价提供了新的技术手段。We used inter-primer binding site PCR （iPBS-PCR） to amplify the relevant fragments from the northwest-plain tree peony Red Hydrangea and central-plain variety Luoyanghong, then the polymorphic fragments were cloned, sequenced and analyzed by relevant bioinformatics software; 12 long terminal repeat （LTR） sequences were obtained from peony LTR retrotransposons. The nucleotide sequence had the highest heterogeneity, deletion mutation was the main behavior, the length of the nucleotide sequences ranged from 313 to 894 bp, and homologous sequences ranged from 31.1% to 65.8%. Cluster analysis of the amino acid sequence and the LTR retrotransposons from different plant LTR amino acid sequences revealed high homology between relevant sequences of some other plants, which suggests transverse transfer between the LTR retrotransposons. Furthermore, we designed primers according to the cloned LTR sequence and analyzed 29 different varieties of peonies using the sequence-specific amplification polymorphism molecular marker method, which showed high polymorphism. LTR sequences from peony using iPBS may be useful and provide a new technology for peony germplasm resource evaluation.

关 键 词：iPBS LTR类反转录转座子 序列分析 SSAP分子标记 

分 类 号：S685.11[农业科学—观赏园艺]