香蕉果实果胶裂解酶基因cDNA克隆及序列分析

利用已报道果胶裂解酶基因的保守序列设计简并引物,进行RT-PCR,得到1个大约1300bp的香蕉果胶裂解酶基因cDNA片段,命名为MA-pl。DNA序列分析表明:MA-pl片段全长1277bp,包含1个882bp的开放读码框(ORF),编码294个氨基酸;其具有所有果胶裂解酶共有的保守区域:钙协调部位(Asp72,Asp74,Asp96,andAsp100)、酶活性位点(Arg152,Pro154,Arg157)及3个重要的结构域(motifI:WVDH,motifII:DGLVDAVMGSTAITVSNNYF,motifIII:LYQRMPRCRHGYFHVVWNDY);MA-pl氨基酸序列与草莓-1、葡萄、草莓-2、拟南芥、苹果、香蕉(banana-1)的相似性分别为85.8%、74.2%、79.7%、78.6%、72.4%和71.4%。

Cloning and sequence of cDNA fragments encoding pectate lyase genes from banana fruit

A 1277 bp cDNA fragment encoding pectate lyase, named as MA-pl, was cloned from banana fruit by using degenerate primers designed with reference to the conserved amino acid sequences of known pectate lyases by RT-PCR. Sequence analysis showed that MA-pl contains an 882 bp ORF, encoding a predicted polypeptide of 294 amino acids. Alignment of the deduced amino acid sequence of MA-pl and other putative pectate lyases showed that MA-pl has an identity of 85.8%,74.2%, 79.7%, 78.6%, 72.4% and 71.4% respectively to pectate lyases from strawberry-1, grape, strawberry-2, arabidopsis thaliana, apple and banana-1. The amino acid residues conserved in all pectate lyase genes involved in Ca2 coordination (Asp72, Asp74, Asp96, and Asp100) and the active site conformation (Arg152, Pro154, Arg157) were present in MA-pl. Three conserved motifs (motif I: WVDH, motif II: DGLVDAVMGCSTAITISNNHL, motif III: LYQRMPRCRHGYFHVVW NDY) characteristic to all pectate lyases were found in MA-pl, suggesting that MA-pl highly likely encodes a pectate lyase in banana.