日本蛇根草查尔酮异构酶基因的克隆及其生物信息学分析

Cloning and Bioinformatics Analysis of CHI Genes in Ophiorrhiza japonica

查尔酮异构酶(CHI)是类黄酮生物合成过程中的关键酶之一,其在类黄酮的合成过程中发挥着非常重要的作用。本研究以日本蛇根草为研究材料,根据其转录组测序结果设计全长引物,通过RT-PCR方法成功克隆得到日本蛇根草查尔酮异构酶基因完整的cDNA序列,并对其进行生物信息学分析。结果表明,该基因序列全长为702 bp,编码233个氨基酸,预测其蛋白分子量为25.018 k D,等电点为4.95,是亲水性蛋白质,不含有信号肽,很可能定位在叶绿体基质中。同时,二级结构分析显示,日本蛇根草CHI的二级结构分别由96个α螺旋、51个延伸链、86个无规则卷曲组成,这与三维结构预测结果相一致。日本蛇根草CHI基因的克隆对于研究植物类黄酮的生物合成及其分子机制具有重要意义,同时也丰富了双子叶植物CHI基因家族的相关研究。

Chalcone Isomerase （CHI） is one of the first enzymes in the flavonoid biosynthetic pathway, and it plays an important role during flavonoid biosynthesis. In this study, we used Ophiorrhiza japonica as materials, and designed the primers according to the result of its transcriptome sequencing. The integrated cDNA of CHI was cloned successfully by RT-PCR method, and then was applied to bioinformatics analyses. These results showed that the full-length CDS of CHI was 702 bp, and encoded the protein which consisted of 233 amino acids. The putative protein molecular weight was 25.018 kD and its isoelectric points were 4.95 CHI of Ophiorrhizajaponiea was a hydrophilic protein, had no signal peptide, and was likely to locate in the chloroplast stroma. Meanwhile, analysis of secondary structure indicated that CHI of Ophiorrhiza japonica contained 96 alpha helixs, 51 extended strands, 86 random coils, and this result was consistent with the prediction of its three dimensional structure. Cloning of CHI from Ophiorrhiza japonica has an important significance for the research of flavonoid biosynthesis and its molecular mechanism in plants, but also enriches the correlation of CHI gene family members in dicotyledones plants.