黄瓜CsGDSL脂解酶基因的克隆与表达分析

Cloning and Expression Analysis of CsGDSL Lipase Gene in Cucumber

GDSL脂解酶是影响角质层发育的重要基因,克隆黄瓜GDSL脂解酶基因,并分析其表达模式,旨在为黄瓜角质层发育及黄瓜果实光泽的研究奠定基础。根据黄瓜基因组数据库中的参考序列,对黄瓜GDSL脂解酶基因进行克隆,并进行生物信息学分析。利用qRT-PCR技术明确该基因在黄瓜各组织部位中的表达情况。以6份光泽性不同的黄瓜为材料,对该基因序列进行克隆,以明确该基因在不同光泽性黄瓜中的作用。对黄瓜GDSL脂解酶基因启动子进行克隆并分析其作用元件。参照黄瓜基因组数据库设计扩增引物进行PCR扩增,获得CsGDSL基因,CDS长度1059 bp,编码352个氨基酸,二级结构以无规卷曲(45.45%)与α-螺旋(33.24%)为主;该基因在进化过程中较为保守,与甜瓜CmGDSL亲缘关系最为紧密;在黄瓜花后3 d的子房中表达量较开花0 d的子房高;在6个品种中CsGDSL基因CDS序列保守;CsGDSL基因与逆境胁迫、激素及光等具有响应。获得了CsGDSL脂解酶基因,明确了其在黄瓜不同组织部位中的表达情况,该基因在不同材料中较为保守,由此推测,黄瓜CsGDSL可能影响黄瓜果皮光泽。

GDSL is an important gene affecting the development of cuticle.This study cloned cucumber GDSL lipase gene and analyzed its expression pattern,in order to lay a foundation for the study of cuticle development and the glassiness of cucumber.The gene sequence was cloned to clarify its role in cucumbers with different glossiness by using six cultivars with different glossiness as materials.The promoter of CsGDSL lipase gene was cloned to analyze its functional elements.According to the reference sequence in cucumber genome database,we cloned the GDSL lipase gene of cucumber and analyzed by bio-informatics.The expression of the gene in different tissue parts of cucumber was confirmed by qRT-PCR technique.The CDS of CsGDSL length 1059 bp which encoded 352 amino acids,and the secondary structure was mainly random curl(45.45%)andα-helix(33.24%).This gene was conservative in the process of evolution and was most closely related to CmGDSL.The expression level was the highest in the male flower of cucumber on the flowering day,and the expression level in the ovary at 3 days after flowering was higher than that at 0 days after flowering.The CDS sequence of CsGDSL gene was conservative in six cultivars.CsGDSL gene was responsive to stress,hormone and light.We obtained the CsGDSL lipase gene in this study,and identified its expression in different tissue parts of cucumber,the gene is relatively conserved in different cultivars,suggesting that CsGDSL may affect cucumber glossiness.