条斑紫菜eIF4A基因的电子克隆与试验验证

Cilico Cloning and Verification of eIF4A Gene from Laver Porphyra yezoensis

以拟南芥eIF4A氨基酸序列为信息探针,在条斑紫菜EST(Expressed sequence tag)数据库中找到高度相似的EST,通过人工序列拼接及RT-PCR确认得到了翻译起始因子4A(Eukaryotic transla-tion initiation factors 4A,eIF4A)基因PyeIF4A的cDNA序列。该cDNA序列含有长1278 bp的完整开放阅读框,编码蛋白(PyeIF4A)分子量47.4 kDa,长425 AA。PyeIF4A与小鼠、非洲爪蟾、水稻和拟南芥等多种动植物的eIF4A具有较高的序列一致性,含有eIF4A的保守基序。

The silico cloning, verification and characterization of eukaryotic translation initiation factors 4 A（eIF4A）genes was conducted in cDNA of laver Porphyra yezoensis （PyelF4A）. The highly similar P. yezoensis ESTs were obtained by Arabidopsis thaliana eIF4A amino acids sequence as a querying probe from dbEST of GenBank and the putative cDNA sequence of PyeIF4A was assembled. Furthermore, the cDNA of PyelF4A was isolated by RT-PCR with two primers designed in accordance with the assembled sequence and was found to contain an 1278 nt of continuous complete open reading frame encoding a polypeptide（PyeIF4A）of 425 amino acids with a calculated molecular weight of 47.4 kDa. Sequence comparison of the PyeIF4A reveals high amino acid sequence identity with eIF4A from other organisms, such as mice Mus musculus , toad Xenopus laevis , rice Oryza sativa and plant Arabidopsis thaliana. The PyeIF4A contained conserved amino acids residue and sequences motifs closely related to function of eIF4A.