银缕梅MADS-box家族基因PsuAP1的克隆、生物信息学表达分析

本研究以珍稀濒危植物银缕梅为材料,利用同源克隆与RACE技术克隆得到一个AP1-like基因的c DNA全长序列,命名为Psu AP1。利用生物信息学方法对它所编码的氨基酸序列进行分析,结果表明,该基因序列全长916 bp,开放阅读框长度为747 bp,编码248个氨基酸,分子质量约28.62 k D,理论等电点为9.84,是一种不稳定亲水蛋白。保守结构域分析表明该基因属于MADS-box家族基因;序列比对分析显示其氨基酸序列与连香树相似性最高,达78.57%,进化树聚类分析也表明Psu AP1与连香树的氨基酸亲缘关系最近。亚细胞定位分析显示该蛋白定位于细胞核。二级结构中同时含有琢-螺旋、延伸链和无规则卷曲结构,分别占比47.18%、11.29%、41.53%。荧光实时定量PCR分析表明,Psu AP1基因在花及果实中均有大量表达,推测其与花的发育以及果实的成熟相关。Psu AP1基因的克隆以及表达分析使银缕梅开花调控机理的研究取得了新的突破,为该基因功能的进一步深入研究提供了实验材料以及理论基础。

Cloning of MADS-box Gene PsuA P1 from Parrotia subaequalis and Bioinformatics Expression Analysis

This study uses the rare and endangered plant Parrotia subaequalis as material,the full length of a AP1-like c DNA was cloned by the technology of homology cloning and RACE,named Psu AP1.Using the bioinformatics method to analyze the amino acid sequence encoded by Psu AP1,the results showed that the c DNA of Psu AP1 is 916 bp,the length of open reading frame(ORF)is 747 bp,encoded 248 predicted amino acids,while the estimated molecular weight and isoelectric point of the putative protein were 28.62 k D and 9.84.Psu AP1 is an unstable hydrophilic protein.Conserved domain analysis indicated that Psu AP1 belonged to the MADS-box family;Sequence alignment analysis showed that the amino acid sequence had the highest similarity with the Cercidiphyllum japonicum,reaching 78.57%.The phylogenetic tree analysis also showed that Psu AP1 was closely related to the C.japonicum.Subcellular localization analysis revealed that Psu AP1 was localized to the nucleus.The secondary structure containsα-helix,extended chain and random coil structure,accounting for 47.18%,11.29%and 41.53%,respectively.Fluorescence real-time quantitative PCR analysis showed that Psu AP1 was abundantly expressed in flowers and fruits,which was predicted to be related to flower development and fruit ripening.The cloning of Psu AP1 gene has made a new breakthrough in the study of flowering regulation mechanism of P.subaequalis,also provided substantial experimental foundation for further studying on the function of Psu AP1.