摘要
根据莱茵衣藻(Chlamydomonas reinhardtii)、Oryza sativa、Chlorella vulgaris以及Me-sostigma viride等真核生物的psbD基因的氨基酸高度保守序列,设计一对简并引物,利用TRIzol试剂提取杜氏盐藻(Dunaliella salina)细胞的总RNA,通过RT-PCR,得到杜氏盐藻cDNA片段的长度大约为1 000bp.该序列的PCR产物经T-A克隆并测序分析以及测序结果推导成氨基酸序列,Blast同源性分析表明所克隆的基因为杜氏盐藻psbD基因,编码杜氏盐藻光系统Ⅱ反应中心D2蛋白,该序列已递交GenBank(GenBank登录号为:DQ074450).编码的氨基酸序列,同源性依次为:Chlamydomonas reinhardtii92%,Nephroselmis olivacea88%,Pinus thunbergii88%,Am-borella trichopoda88%,Chlorella vulgaris88%.通过密码子偏爱性分析表明,psbD基因存在明显的密码子偏爱性,其(A+T)含量明显高于(G+C)含量.
One pair of degenerate primers was designed according to conserved motifs ot the psbD （encoding PhotosystemⅡreactor center proteins D2 ） of Chlarnydornonas refnhardtii , Oryza sativa , Chlorella vulgaris and Mesostigrna viride. And a total RNA of Dunaliella salina （D. salina） was extracted with TRIzol reagent. A cDNA fragment, 1000bp in length, from green algal D. salina was obtained through RT-PCR method. The resulting PCR products were cloned into T-vector and screened to determine their sequence. Homologous analysis of the deduced amino acid sequence was performed by BLAST and subsequently compared to that of other species, it is shown that the sequence cloned is a cDNA fragments of psbD from D. salina, the sequence has submitted to GenBank（GenBank accession number： DQ074450）. In addition, homology analysis of psbD shows that Dunaliella salina are high identity with the following species： Chlamydornonas reinhardtii 92%, Nephroselrnis olfvacea 88%, Pinus thunbergii 88%, Amborella trichopoda 88%, and Chlorella vulgaris 88%. Also, the analysis of codon bias reveals that the codon usage of psbD gene of D. salina is apparently biased, and the content of （A＋T） is obviously higher than that of （G＋C）, in the meanwhile, the numbers of Lys in the psbD gene is 3, and this property will provide the theoretical basis of isolation, purification and study of the proteins of D2.
出处
《华中师范大学学报（自然科学版）》
CAS
CSCD
2007年第3期431-436,共6页
Journal of Central China Normal University：Natural Sciences
基金
国家自然科学基金资助项目(30600006).
