甘蓝型油菜oleosin基因片段的克隆及序列分析

Cloning and Sequence Analysis of an Oleosin Gene Fragment from Brassica napus

以油菜(Brassica napus)基因组DNA为模板,通过设计一对特异性引物,利用多聚酶链式反应(PCR)扩增获得oleosin基因片段。将其克隆到E.coli质粒上进行序列分析。结果表明,该基因由878个碱基组成,含一个内含子,编码193个氨基酸。不计附加的18bp凝血酶切割序列,与已报道的序列相比较,核苷酸序列及推导出的氨基酸序列的同源性分别为79.20%和91.21%。推测的氨基酸序列构成的蛋白质具有oleosin的基本结构特点,包含3个结构区域:两性N末端区(anamphipathicN-terminalre鄄gion),中心疏水区(acentralhydrophobicdomain),两性C-末端区(anamphipathicC-terminaldomain)。

The oleosin gene fragment was amplified by means of PCR using genomic DNA of Brassica napus as template and a pair of specific oligonucleotides at 5'- and 3'-end as primer, and cloned into E.coli vector. Results show that the DNA sequence consists of 878 bp and encodes 193 amino acid residues deduced from DNA sequence, which have an intron. Except for the extra thrombin cleavage site of 18 bp, comparison with previously published sequence showed 79.20% homologies in nucleotide sequence and 91.21% in amino acid sequence respectively. The deduced protein has the same structural regions, an amphipathic N-terminal region, a central hydrophobic domain, and an amphipathic C-terminal domain.