盐芥EsABCG25的克隆及其表达分析

doi:10.13560/j.cnki.biotech.bull.1985.2018-0020

摘要/Abstract

摘要： 旨在揭示盐芥（Eutrema salsugineum）ABCG25的结构及其对干旱、盐害以及温度等胁迫的响应。以山东生态型盐芥作为材料,通过电子克隆以及RT-PCR技术,获得一个腺苷三磷酸结合盒转运蛋白G（ABCG）家族成员基因的全长cDNA序列,命名为EsABCG25,GenBank登录号为KY111263。EsABCG25全长2 154 bp,含1 983 bp的开放阅读框,编码一个含有660氨基酸的蛋白分子。该基因定位于盐芥第5染色体,含有4个外显子和3个内含子。EsABCG25蛋白具有一个核苷酸结合结构域（NBD）和一个跨膜结构域（TMD）,与来源于同科植物芜菁的ABCG25亲缘关系最近。对高通量测序数据分析表明该基因在盐芥的茎部表达最为丰富,在莲座叶中表达丰度最低,且在不同组织中均有可变剪接事件。Real-time PCR结果显示,该基因表达受多种逆境影响,尤其受到ABA的明显诱导。EsABCG25具有ABCG家族的典型序列特征;该基因在盐芥不同组织中的表达丰度存在差异,表达受到多种逆境的诱导,与盐芥的抗逆性关系密切。

关键词: 盐芥, ABCG25, 生物信息学, Real-time PCR, 表达分析

Abstract: This work is aimed to reveal structure of ABCG25 and explore its response to drought,salinity and temperature stresses in Eutrema salsugineum. A full-length cDNA of member gene in ATP-binding cassette transporters（ABCG）family was cloned from E. salsugineum（Shandong ecotype）using silicon cloning and RT-PCR methods,and named as EsABCG25 with GenBank accession number of KY111263. The full length of EsABCG25 was 2154 bp and it contained a complete ORF with 1983 bp and encoded 660 amino acids. EsABCG25 was located at chromosome 5,with 4 exons and 3 introns. EsABCG25 had a putative nucleotide binding domain（NMD）and a putative transmembrane domain（TMD）. Alignment and phylogenetic analysis found that EsABCG25 was the closest to the ABCG25 from Brassica rapa. High-throughput sequencing indicated that EsABCG25 reached the highest expression in the stem of E. salsugineum and the lowest expression in rosette leaves,and alternative splicing events were detected in all tissues. Real-time PCR assay indicated that EsABCG25 was induced by many kinds of stresses,especially ABA stress. From above,EsABCG25 has typical characteristics of ABCG gene family. Its expression variedat different tissues,was induced by stress,and may be closely associated with the stress tolerance of E. salsugineum.

Key words: Eutrema salsugineum, ABCG25, bioinformatics, real-time PCR, expression analysis

松布尔巴图, 陈悦, 陈宁美, 唐帅, 徐小静. 盐芥EsABCG25的克隆及其表达分析[J]. 生物技术通报, 2018, 34(7): 108-118.

Songbuerbatu CHEN Yue CHEN Ning-mei TANG Shuai XU Xiao-jing. Cloning and Expression of EsABCG25 from Eutrema salsugineum[J]. Biotechnology Bulletin, 2018, 34(7): 108-118.

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