半套式PCR技术在植物基因克隆中的应用  

Utilization of Half Nested PCR in Plant Gene Cloning

作　　者：杨明挚[1,2] 陈善娜[1,2] 鄢波 黄兴奇[1,2] 刘继梅[1,2] 

机构地区：[1]云南大学生物系 [2]云南省农科院生物技术所

出　　处：《云南大学学报（自然科学版）》1998年第5期377-379,共3页Journal of Yunnan University(Natural Sciences Edition)

基　　金：国家自然科学基金;云南省应用基础研究基金;云南省教委基金;云南省农业生物技术重点实验室开放基金

摘　　要：常规ＰＣＲ方法在分离克隆已知序列的基因中发挥着重要的作用，但要求目的基因片段与引物间的同源性几乎是１００％，一旦引物序列与目的基因间存在有差异，便往往会导致扩增的特异性不强，扩增效率不高，给克隆带来很大的困难．在常规ＰＣＲ基础上，若所分离的基因在不同植物间存在保守序列，依据这一序列再合成一对引物，进行半套式ＰＣＲ则能大大提高扩增的特异性及扩增效率．对于较长的基因片段而言还能同时完成亚克隆．本文依据南瓜ＧＰＡＴ（甘油－３－磷酸酰基转移酶）基因ｃＤＮＡ序列及比较拟南芥菜。The normal PCR playing a very important role in cloning of a sequence known gene.But high homologous (almost 100%) between primers and target gene sequence must be required.Otherwise,low efficiency and specifity of amplification may be.If homologeous regions exist among different plants in this gene,then prepare another pair of primers from these regions,using half nested PCR,the amplification efficiency and specifity will be greatly improved.Here we indicate the applification of half nested PCR technique in plant gene cloning by isolation and cloning of figleaf gourd ( cucurbita ficifolia ) and watermelon ( Citrullus lanatus ) glycerol 3 phosphate acyltransferase (GPAT) gene using a pair of primers from squash and another pair of primers from the homologeous region of this gene as example.

关 键 词：半套式PCR 基因克隆 植物 

分 类 号：Q781[生物学—分子生物学]