伽师瓜和86-1甜瓜几丁质酶基因的克隆与序列分析

Cloning and Sequence Analysis of Chitinase Gene from Jiashi Melon and 86-1 Melon

以新疆主产不同耐藏性的甜瓜品种伽师瓜(耐藏性好)和86-1甜瓜(新密11号,耐藏性差)为试料,利用甜瓜基因组辅助设计引物克隆几丁质酶基因的c DNA序列,分别对其编码序列(CDS)及编码氨基酸进行相关生物信息学的预测分析,以期通过研究二者之间基因的差异揭示伽师瓜和86-1甜瓜采后不同耐藏性的机制。结果表明,获得的伽师瓜几丁质酶基因(Jia Shi Melon chitinase,JSMC2),NCBI检索号为KT921406;86-1甜瓜几丁质酶基因(Xin Mi-11 melon chitinase,XMMC),NCBI检索号为KU236388。JSMC2和XMMC序列长度均为1 011 bp,包含完整的开放阅读框,编码336个氨基酸,与黄瓜几丁质酶基因具有高度的同源性。蛋白结构域预测分析表明,甜瓜几丁质酶蛋白包含GH18_hevamine_Xip I_class_Ⅲ、Chi1、GH18_chitinase-like super family、Glyco-hydro-18和Glyco_18保守结构域,可催化几丁质水解成N-乙酰葡糖胺,分解真菌细胞的细胞壁,抵御病原真菌的侵染。

Two melon varieties with different storage properties from Xinjiang： Jiashi melon（good storage quality） and86-1 melon（Xinmi 11, bad storage quality） were used as the experimental materials. The c DNA of chitinase genes were cloned from Jiashi melon and 86-1 by using genome of melon to design primer, cooding sequence（CDS） and encoded amino acid of chitinase were also predicted by bioinformatics analysis. The purpose of this paper was to reveal the different storage resistance mechanism of post-harvest Jiashi melon and 86-1 melon by investigating genetic differences between the two melons. The results showed that, the NCBI accession number of chitinase gene of Jiashi melon（Jia Shi Melon chitinase, JSMC2） was KT921406, and the NCBI accession number of chitinase gene of86-1 melon（Xin Mi-11 melon chitinase, XMMC）, was KU236388. Both JSMC2 and XMMC sequence length were1 011 bp, which contained a complete open reading frame, encoded 336 amino acids, and had a high degree of homology with cucumber chitinase. The predicted analysis of protein domain showed that JSMC2 and XMMC contained GH18_hevamine_Xip I_class_ Ⅲ, Chi1, GH18_chitinase-like super family, Glyco-hydro-18 and Glyco_18, whichcould catalyze the hydrolysis chitin to N-acetylglucosamine, decompose fungal cell wall and resist fungal pathogen infection.