棕色棉F3'H-羟化酶基因的克隆与生物信息学分析  

Cloning and Bioinformatics Analysis of Flavonoid 3′-Hydroxylase Gene GhF3′Hin Brown Cotton(Gossypium hirsutum L.) Fibers

作　　者：梁明炜[1] 刘海峰 肖向文[1] 陆雪莹[1] 李艳红[1] 宋武 鲁春芳[1] 李晓波[1] 

机构地区：[1]中国科学院新疆理化技术研究所新疆植物资源化学重点实验室,乌鲁木齐830011 [2]中国彩棉(集团)股份有限公司,乌鲁木齐830016

出　　处：《生物技术通报》2011年第11期199-206,共8页Biotechnology Bulletin

基　　金：中国科学院新疆理化技术研究所博士启动资金项目;中国科学院"西部之光"人才培养计划项目(RCPY200802);转基因生物新品种培育重大专项项目(2009ZX08005-027B)

摘　　要：根据葡萄的类黄酮3'-羟化酶(F3'H)基因全长cDNA序列Blast所得棉花的EST序列设计引物,以开花后16 d(DPA16)的新彩棉5号(XC-5)纤维为材料,利用RACE和RT-PCR技术分离得到了2个类黄酮3'-羟化酶基因cDNA序列,此2个序列编码区完全相同,仅在3'UTR区存在片段长短的差异,推测可能是基因转录后加工方式不同所造成。克隆所获得的棉花F3'H基因编码区全长1 533 bp,编码510个氨基酸,氨基酸序列分析预测表明,该基因所编码蛋白含有一个跨膜结构域,是一种分泌蛋白,定位于内质网上,并含有一段与细胞色素P450功能区相匹配的保守功能域;序列比对结果表明,棉花F3'H基因与其他多个物种的F3'H基因在氨基酸序列上有较高的同源性;聚类分析结果表明,棉花F3'H蛋白与双子叶植物大豆的F3'H亲缘关系较为接近,而与单子叶植物高粱等作物则较远。Two full-length cDNA sequences encoding flavonoid 3′-hydroxylase gene（F3′H） were cloned from brown-colored cotton（Gossypium hirsutum L.） cultivar ＂XC-5＂ fibers of 16 days post anthesis（DPA） by rapid amplification of cDNA ends（RACE） and RT-PCR technique using specific primers based on the EST sequences from the blast results with the full-length cDNA of F3′H in Vitis vinifera and were named as GhF3′H-1 and GhF3′H-2（GenBank accession： HM598123,HM598124）.The two cDNA sequences contain an identical opening reading frame（ORF） of 1 533 bp in length,encoding a protein of 510 amino acids.There are few differences between the two sequences except for the sequence length of the 3′untranslational region（UTR）,which might result from the difference of post-transcriptional processing.The bioinformatics analyses showed that,the GhF3′H protein,holding with a membrane spanning domain,are located on endoplasmic reticulum and belongs to secretory proteins;in addition,it contains a conservative functional domain which matches the cytochrome P450 binding domain;the GhF3′H gene shows the highly homogeneity at amino acid level in comparison to the F3′H genes of several other higher plant species;the phylogenetic analysis of F3′H genes from different species reveals that the GhF3′H has closer relationship with the F3′H from Glycine max than from other plant species.

关 键 词：类黄酮3'-羟化酶 棕色棉 RACE 序列分析 

分 类 号：S562[农业科学—作物学]