摘要
为了解刺槐(Robinia pseudoacacia)Kuntiz型胰蛋白酶抑制剂基因(RpKTI)的功能,本试验提取刺槐叶片中RNA,采用RACE PCR技术克隆RpKTI基因序列全长,并对其进行生物信息学分析及表达分析。结果表明,RpKTI基因的开放阅读框(ORF)长度为633 bp,共编码210个氨基酸,分子式为C_(1039)H_(1586)N_(258)O_(307)S_(5),是一个稳定的酸性疏水蛋白;在氨基酸序列的N端存在一个跨膜区,且在此跨膜区内含有信号肽;具有11个磷酸化位点及1个N-糖基化位点;二、三级结构均以延伸链及无规则卷曲为主。同源性及进化树分析表明,RpKTI基因属于STI超级家族,与蒺藜苜蓿(Medicago truncatula)的KTI基因亲缘关系更近。通过实时荧光定量(qRT-PCR)分析发现RpKTI基因在铅镉混合胁迫3 d时,叶中的表达量最高,推测此基因可能在重金属铅镉胁迫下发挥重要作用。本研究为后期RpKTI基因的功能验证及应用研究奠定了理论基础。
In order to understand the function of Kuntiz trypsin inhibitor gene RpKTI of Robinia pseudoacacia, the RNA was extracted from R. pseudoacacia leaves, and the full length of RpKTI gene sequence was cloned by RACE PCR, then its bioinformatics analysis and expression analysis were carried out. The results showed that the open reading frame(ORF) length of RpKTI gene was 633 bp, encoding 210 amino acids, and the molecular formula wasC_(1039)H_(1586)N_(258)O_(307)S_(5),which was a stable acidic hydrophobic protein. There was a transmembrane region at the N-terminal of the amino acid sequence, and there was a signal peptide in this transmembrane region. There were 11 phosphorylation sites and one N-glycosylation site. The secondary structure and tertiary structure were mainly extended strand and random coil. Through homology and phylogenetic tree analysis, RpKTI gene belonged to STI superfamily, which was more closely related to the KTI gene of Medicago truncatula. qRT-PCR analysis showed that the expression of RpKTI gene was the highest in leaves under lead and cadmium combined stress for 3 days. It suggested that this gene might play an important role in the stress of heavy metal lead and cadmium. This study laid theoretical foundation for the functional verification and application research of RpKTI gene in the later stage.
作者
张晓静
董启迪
王霞
张晓娜
周建
Zhang Xiaojing;Dong Qidi;Wang Xia;Zhang Xiaona;Zhou Jian(Schoo of Horiculture and Landscape Architecture,Henan Institute of Seience and Technology,Xinriang 453003,China;Changyun Forestry Bureau,Changyuan 453400,China;Henan Horticultural Plant Resource Utilization and Germplasm Innovation Engineering Research Center,Xinxiang 453003,China)
出处
《山东农业科学》
北大核心
2022年第6期10-19,共10页
Shandong Agricultural Sciences
基金
河南省科技攻关项目“刺槐修复铅镉污染土壤的双向调控关键技术研究及应用”(202102110053)。
关键词
刺槐
RpKTI基因
基因克隆
生物信息学分析
表达分析
Robinia pseudoacacia
RpKTI gene
Gene cloning
Bioinformatics analysis
Expression analysis
