巴夫杜氏藻β-肌动蛋白基因的克隆和分析

目的]获得巴夫杜氏藻β-肌动蛋白基因cDNA全长序列。方法]以巴夫杜氏藻cDNA为模板,采用简并引物进行PCR扩增,获得533bp特异cDNA片段。在此基础上,设计特异引物,采用5’-Genome Walking和3’-RACE的方法,获得基因的5’-端DNA序列和3’-端cDNA序列,进而获得β-肌动蛋白基因cDNA全长序列。结果]获得了巴夫杜氏藻β-肌动蛋白基因的特异cDNA片段、5’-端DNA和3’-端cDNA片段。经拼接后,扩增出全长cDNA。β-肌动蛋白基因cDNA全长1754bp,包括1137bp的开放读码框和617bp的3’-非翻译区序列。氨基酸序列相似性分析发现,巴夫杜氏藻β-肌动蛋白氨基酸序列与杜氏盐藻、莱茵衣藻等的同源性较高。系统发育分析表明,巴夫杜氏藻β-肌动蛋白与杜氏盐藻的相似性最高。结论]首次获得了巴夫杜氏藻β-肌动蛋白基因cDNA全长序列,并发现巴夫杜氏藻β-肌动蛋白基因非常保守。

Cloning and Characterization of Beta-actin Gene in Dunaliella parva

Objective] The aim was to obtain the full-length cDNA sequence of Dunaliella parva β-actin gene.Method] Based on the highly conserved amino acid regions of known β-actin,a pair of degenerate primers was synthesized to amplify 533 bp cDNA sequences in Dunaliella parva.Then,the 5' genomic DNA and 3' cDNA sequences were obtained by Genome walking and 3'-RACE technology based on the obtained sequence.According to the sequences of the 5'-termini and 3'-termini,specific primers were synthesized to obtain the full-length cDNA.Result] The full-length β-actin cDNA included 1 137 bp open reading frame(ORF),617 bp of 3' noncoding region.Similarity analysis indicated that the highest similarity was found between Dunaliella parva and Dunaliella salina.The Dunaliella parva β-actin also showed wide similarity with other algae.Conclusion] The full-length cDNA sequence of D.parva was firstly obtained,which was highly conserved.