摘要
根据真核生物莱茵衣藻(Chlamydomonasreinhardtii)、Chlamydomonasmoewusii、Chlorellavulgaris以及Mesostigmaviride的psaB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(Dunaliellasalina)细胞的总RNA,通过RT-PCR,得到的一段长为1·8kb左右的cDNA片段。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进行同源性比较,表明所克隆的1815bp序列为杜氏盐藻psaBcDNA片段,GenBank收录号为AY820754。根据已经得到的psaB序列推导成氨基酸序列与一些已知物种的psaB基因相比较,同源性分别为Chlamydomonasreinhardtii92%,Chlamydomonasmoewusii91%,Chlorellavulgaris86%,Mesostigmaviride85%,Physcomitrellapatenssubsp.Patens85%,Nephroselmisolivacea84%。据此可推断本实验中所克隆的序列为杜氏盐藻psaBcDNA序列。
One pair of degenerate primer was designed according to conserved motifs of the psaB(A2 subunit of photosystem Ⅰ) of Chlamydomonas reinhardtii, Chlamydomonas moewusii, Chlorella vulgaris and Mesostigma viride, and a total RNA of Dunaliella salina (D.salina) was extracted with TRIzol reagent. A cDNA fragment, about 1.8kb in length, from green algal D.salina was obtained through RT-PCR method. The resulting PCR product was cloned into T-vector and screened to determine its sequence. Homologous analysis of the deduced amino acid sequence was performed by BLAST and subsequently compared with GenBank data. The obtained cDNA sequence was 1815 bp long, which encodes 605 amino acids (GenBank accession number: AY820754). The sequence shared high homologue with the following psaB: Chlamydomonas reinhardtii 92%, Chlamydomonas moewusii 91%,Chlorella vulgaris 86%, Mesostigma viride 85%, Physcomitrella patens subsp. Patens 85% and Nephroselmis olivacea 84%. It can be concluded that the cloned sequence is psaB cDNA fragment from D.salina.
出处
《生物工程学报》
CAS
CSCD
北大核心
2005年第4期642-645,共4页
Chinese Journal of Biotechnology
基金
国家自然科学基金资助项目(No.30270031
30300208)
河南省医学科技创新人才工程项目(No.2001006)~~