柑橘类植物GPAT基因片段克隆和SNP分析  

Cloning of Partial Sequence of GPAT Gene in Citrus and Its SNP Analysis

作　　者：吴波[1] 刘勇[2] 

机构地区：[1]江西中医学院江西省中药种质资源工程技术研究中心,江西南昌330004 [2]江西农业大学农学院,江西南昌330045

出　　处：《江西农业大学学报》2010年第1期51-56,共6页Acta Agriculturae Universitatis Jiangxiensis

基　　金：江西省自然科学基金资助项目(2009GZN0024);产学研结合培养高素质复合型中药人才模式创新研究与实践项目(教高函[2007]29号);江西农业大学博士启动基金项目(1791)

摘　　要：甘油-3-磷酸酰基转移酶基因是与植物抗冷性强弱密切相关的基因。根据已报道的各种植物GPAT基因的氨基酸序列上前后两个保守区设计1对简并引物,采用RT-PCR技术,分别从琯溪蜜柚、马家柚、莽山野橘等8种柑橘类植物中克隆得到315bp的GPAT基因片段。使用DNA Star5.0软件对8个片段序列进行同源性比较,8个片段的核苷酸同源性高达97.8%,与其它植物GPAT基因核苷酸序列的同源性都在72.5%以上;经过氨基酸序列推导,每个GPAT基因片段分别编码105个氨基酸,8个片段的氨基酸同源性为95.2%,与其它植物GPAT基因的氨基酸序列同源性在68.9%以上。试验克隆所得8种柑橘类植物GPAT基因片段序列存在明显的单核苷酸变异,在14个核苷酸位点存在SNPs(single nucleotide polymorphisms),导致6个氨基酸位点发生变异,多态性频率为1SNP/22.5bp,核苷酸变异度为4.45%,氨基酸变异度为5.71%。采用NJ聚类法对克隆片段的核苷酸序列进行了遗传多样性分析。GPAT gene is an important gene with cold resistance of plants.A pair of degenergte primers were designed according to the conserve regions of GPAT gene of many kinds of plants reported in Genebank.Eight cDNA documents of GPAT gene in Citrus maxima cv.Guanximiyou,Citrus maxima cv.Majiayou,Citrus mangshanensis,Citrus limon,Citrus sinensis.Xingguotiancheng,Citrus maxima cv.Xinmuyou,Poncirus trifoliate and Citrus maxima cv.Lingnanshatianyou were cloned by using RT-PCR technology.The biology program of DNAStar5.0 was used to analyze the eight document sequences.The best nucleotide similarity was 97.8% among the eight gene documents and the similarity was above 72.5% with other plants＇GPAT gene.Every nucleotide sequence encoded 105 amino acids according to derivation.The best amino acid similarity was 95.2% among the eight gene documents and the similarity was above 68.9% with other plants＇GPAT gene.Fourteen variational sites of single nucleotides were discovered in the eight gene documents.Six amino acid sites transformed because of the variation of fourteen single nucleotides.Accoring to the calculation result,the polymorphism frequence was 1SNP/22.5 bp,the nucleotide and amino acid degree of the variation of nucleotide and amino acids were 4.45% and 5.71%.The genetic diversity of cloning documents was analyzed by using the clustering procedure of NJ（Neighbor-Joining）.

关 键 词：柑橘类植物 GPAT 基因克隆 SNP 序列分析 

分 类 号：Q785[生物学—分子生物学]