水稻基因组MSAP指纹图谱构建及DNA甲基化修饰位点分离与鉴定  

Construction of rice genomic methylation-sensitive amplification polymorphism fingerprint and characterization of DNA methylated sites

作　　者：张勇[1,2] 邓科君[1] 张韬[1] 杨足君[1] 彭金华[1] 周建平[1] 任正隆[1,2] 

机构地区：[1]电子科技大学生命科学与技术学院,成都610054 [2]四川农业大学植物遗传育种省级重点实验室,雅安625014

出　　处：《高技术通讯》2009年第9期983-990,共8页Chinese High Technology Letters

基　　金：国家自然科学基金(30671136、30730065);国家博士后科学基金(20070411158);电子科技大学青年基金资助项目。

摘　　要：采用EcoR Ⅰ和HpaⅡ/Msp Ⅰ双酶切建立了适合于水稻基因组的甲基化敏感扩增多态性(MSAP)分析体系,在全基因组水平检测了水稻DNA甲基化修饰位点。以12对MSAP引物进行选择性扩增,共检测到甲基化修饰位点120个,'CCGG/GGCC'位点甲基化修饰比例为20.17%。对部分水稻基因组甲基化修饰位点进行回收,最终分离了55条存在甲基化位点变异的DNA序列,通过BLAST比对分析将其联配到水稻基因组序列上。分析表明,这些甲基化修饰位点主要集中于基因启动子区(47%)和第一外显子区(22%),在其侧翼序列中存在类似'CpG island'典型序列特征的'CpG'二核苷酸成簇富集区。在此基础上,对应用MSAP技术分离水稻基因组DNA甲基化修饰位点的有效性以及水稻基因组序列中'CpG island'类似序列分布特征和生物意义进行了讨论。In this study, the double digestion of EcoR Ⅰ and Hpa Ⅱ/Msp Ⅰ was used to construct the high-density rice genomic methylation-seusitive amplification polymorphism （MSAP） fingerprint, and the methylated sites were detected at the whole genome level. By using twelve pairs selective primers, a total of 120 methylated sites were detected, which represented that the ratio of methylation modification at the ＇ CCGG/GGCC＇ site in rice genome was 20.17 %. Some of methy- lated sites were extracted and sequenced. Finally, 55 fragments with methylafion modification were characterized and located to rice genome sequence data through the basic local alignment search tool （BLAST）. The further sequence analysis showed that the methylated sites were mainly distributed in the gene promoter region （47 % ） and the first extron region （22%）. The region of CpG rich cluster with typical CpG island sequence features was also detected in the flanking sequence of rice methylated sites. Based on these experimental evidences, the effectiveness to isolate rice methylated site by MSAP and the distribution features of the CpG island-like sequence in rice genome were discussed deeply in the present paper.

关 键 词：水稻 表观遗传 DNA甲基化 甲基化敏感扩增多态性(MSAP) CPG岛 

分 类 号：S511[农业科学—作物学]