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以地黄(Rehmannia glutinosa)叶片c DNA为模板,克隆地黄扩展蛋白基因,获得一条972 bp的c DNA序列,包含了762 bp的开放阅读框(ORF),命名为Rg EXPA10,Gen Bank登录号为KF011918,编码253个氨基酸。以基因组DNA为模板,扩增出一条1 207 bp的序列,测序结果显示包含3个外显子和2个内含子,Gen Bank登录号为KF011919。应用生物信息学方法分析其氨基酸序列结构特征,结果显示,Rg EXPA10包含DPBB_1和Pollen_allerg_1结构域,有26氨基酸的核定位序列和19氨基酸的跨膜结构域。进化分析发现在拟南芥26个α扩展蛋白中,Rg EXPA10与At EXPA8的同源性最高。在22个植物的同源蛋白中,地黄Rg EXPA10与番茄扩展蛋白的亲缘关系最近。以实时荧光定量PCR(q RT-PCR)检测Rg EXPA10基因的表达量,发现Rg EXPA10在地黄未展开的幼嫩叶片里表达最强,其次为膨大中的块根,在衰老的叶片中表达最弱。出苗后60和90天时,在地黄块根中的表达较强。在连作、盐和渍水3种逆境胁迫下,地黄叶片内Rg EXPA10的表达量均显著降低。为研究Rg EXPA10在地黄发育和逆境胁迫中的分子功能奠定基础。
The cDNA of Rehmannia glutinosa was amplified by cloning Rehmannia glutinosa leaf cDNA, and a 972 bp cDNA sequence was cloned, containing a 762 bp open reading frame (ORF) designated as Rg EXPA10 and Gen Bank accession number KF011918 , Encoding 253 amino acids. A genomic DNA was used as a template to amplify a 1 207 bp sequence. The sequencing result showed that it contained three exons and two introns, and Gen Bank accession number was KF011919. Bioinformatics analysis of its amino acid sequence structural features, the results showed that Rg EXPA10 contains DPBB_1 and Pollen_allerg_1 domain, a 26 amino acid nuclear locus and 19 amino acid transmembrane domain. Phylogenetic analysis showed that Rg EXPA10 had the highest homology with At EXPA8 in 26 Arabidopsis α-expansins. Of 22 plant homologues, Rehmannia glutinosa Rg EXPA10 is most closely related to tomato extende. The expression of Rg-EXPA10 gene was detected by q RT-PCR. The results showed that Rg-EXPA10 was the most expressed in young leaves of Rehmannia glutinosa, followed by tuberous roots and the weakest in aged leaves . At 60 and 90 days after emergence, the expression in Rehmannia glutinosa roots was stronger. Under continuous cropping stress, salt stress and waterlogging stress, the expression of Rg EXPA10 in Rehmannia glutinosa Liboschi decreased significantly. It laid the foundation for studying the molecular function of Rg EXPA10 in Rehmannia glutinosa development and stress.