菊花几丁质酶基因ChCHI的克隆及表达分析

根据GenBank发布的几丁质酶基因的保守序列设计简并引物,采用RT-PCR和RACE技术,克隆到菊花1个几丁质酶基因ChCHI(GenBank登录号为KX881373)。ChCHI基因cDNA全长为1 385 bp,包含960 bp开放阅读框,编码319个氨基酸。ChCHI属于亲水性蛋白,相对分子质量约为35.5 k D,等电点为6.38,为稳定蛋白,二级结构预测表明该蛋白以α螺旋和无规则卷曲为主。蛋白序列比对和进化树分析表明,ChCHI基因编码的蛋白属于糖苷水解酶19家族几丁质酶,与薇甘菊(ACO25187.1)几丁质酶蛋白的亲缘关系最近,序列一致性为87%。qRT-PCR分析结果显示,SA处理可以明显提高贡菊叶片中ChCHI的表达量。菊花ChCHI在对抗环境胁迫的分子调控中起重要作用。

Molecular cloning and expression analysis of a chitinase gene ChCHI from Chrysanthemum morifolium

According to the conserved sequences of chitinase gene,the chitinase gene ChCHI was isolated from leaf of Chrysanthemum morifolium using RT-PCR and RACE method (The accession number in GenBank:KX881373).The full-length cDNA of CHCHI was 1385 bp,containing a 960 bp open reading frame which encodes 319 amino acids.Encoding product of ChCHI gene was a kind of hydrophilic and stable protein with a molecular weight of 33.5 kD and isoelectric point of 6.38.Prediction of secondary structure indicated that the protein was mainly composed of alpha helix and random coil.Sequence alignment and phylogenetic analysis showed that the amino acid sequence of ChCHI belongs to family 19 of glycoside hydrolase.CHCHI was highly identified with the chitinase protein from Mikania micrantha (ACO25187.1) with the similarity of 87％.The results of qRT-PCR analysis showed that the expression level of the ChCHI gene in leaves of Chrysanthemum morifolium was significantly increased under the SA treatments.The results implied that ChCHI gene may play an important role in the molecular regulation of resistance to environmental stress in chrysanthemum morifolium.