茶树亲环素基因cDNA全长的分析鉴定与原核表达

通过对茶树新梢cDNA文库大量随机测序，获得了一个编码亲环素的全长基因，在GenBank登录，登录号为DQ904327。茶树亲环素基因cDNA全长949 bp，其中开放阅读框全长495 bp，编码蛋白质含有164个氨基酸，分子量约为17.47 kD，等电点约为8.54，它具备5’端非编码区的“CAAT”标志及3’端非编码区的”AATAAA”poly-A加尾信号。其推测的氨基酸序列经与26条其他生物亲环素蛋白氨基酸序列进行CLUSTAL W多序列联配并以Neighbor-Joining法进行进化树构建后，发现与水稻和小麦的相似性较高，达到85%以上。根据亲环素基因开放阅读框序列设计引物，构建了原核表达载体pET/Csin-Cyp，并在大肠杆菌BL21(DE3)中成功诱导出了一个分子量为23 kD的亲环素融合蛋白。

Molecular Identification, Bioinformatic Analysis and Prokaryotic Expression of the Cyclophilin Gene Full-length cDNA from Tea Plant (Camellia sinensis)

A cDNA clone, encoding cyclophilin, obtained by random sequencing of young shoot cDNA library from tea plant (Camellia sinensis). The full-length cDNA of the cyclophilin gene was 949 bp(GenBank accession No. DQ904327), containing a putative ORF of 495 bp, encoding 164 amino acids and the predicted MW were 17.47 kD and pI was 8.54, respectively. A “CAAT” signal in 5′ untranslated region and a polyadenylated signal of “AATAA”poly-A in 3′untranslated regions of cyclophilin mRNAs were found. The presumed amino acid sequences of tea plant were aligned with those of other 26 organisms through CLUSTAL W. The phylogenetic analysis based on the Neighbor-Joining method showed the similarity was greater than 85% between cyclophilin genes of tea plant and Oryza sativa (japonica cultivar-group), Solanum tuberosum, Triticum aestivum etc. Primers were designed on the open reading frame of the cyclephilin gene of tea plant to construct the expressive vector pET/Csin-Cyp. A recombinant protein about 23 kD in the Escherichia coli BL21 (DE3) was induced.