擎天凤梨花色素合成关键基因CHS、F3'H和DFR的克隆及表达分析

Cloning and Expression Analysis of Guzmania Anthocyanin Biosynthesis Key Genes CHS, F3'H, and DFR

擎天凤梨‘Ostsra’是一种观赏红色苞片为主的高档盆栽花卉。一般认为植物苞片的呈色物质除叶绿素外多来源于花色素,而CHS、F3'H和DFR是在花色素的合成过程中的关键酶。本研究采用采用简并引物及同源克隆法获得了881 bp的CHS基因,编码276个氨基酸的序列,GenBank登录号为KX364270;采用cDNA全长文库构建、EST批量测序及目标单克隆序列测通法,获得了F3'H和DFR基因。F3'H基因长1 176 bp,可编码325个氨基酸序列,GenBank登录号为KX364271;DFR基因长1 209 bp,可编码167个氨基酸的序列,GenBank登录号为KX364272。采用实时定量PCR法检测CHS、F3'H和DFR在‘Ostsra’苞片呈色过程中的表达变化,结果表明,3个基因在苞片变色过程中(绿苞-半红苞-全红苞)都呈现出递增的趋势,即随着苞片颜色的变深,3基因的表达量逐渐增加,全红状态时,表达量最高,而作为对照的绿叶,表达水平都是最低的。

Guzmania ＇Ostsra＇ is high-grade potted flowers with pretty red bracts. Generally, we considered that pigment of plant bracts comes from anthocyanin besides chlorophyll. CHS, F3＇H and DFR are key enzymes in anthocyanin synthesis pathway. In this study, by using degenerate primers and homologous cloning methods, a 881 bp CHS gene fragment was obtained, which coding a sequence with 276 amino acids（GenBank accession： KX364270）;Using full-length cDNA library construction, EST sequencing batch and the target monoclonal whole sequencing methods, we obtained F3＇H and DFR gene. F3＇H gene had 1 176 bp, and encoding a 325 amino acid sequence（GenBank accession： KX364271）. DFR gene had 1 209 bp, and encoding a 167 amino acid sequence（GenBank accession： KX364272）; Through using real-time quantitative PCR to detect CHS, F3＇H and DFR expression level of different coloration bract in ＇Ostsra＇, the results showed that expression level of these three genes gradually increased with deepening red during bracts coloration process（green bract-half red bract-full red bract）. When bract appeared full red, these genes had the highest expression level. However, green leaves as a control, their expression level always the lowest.