【摘要】 目的:克隆杜氏盐藻1,5 -二磷酸核酮糖羧化酶/加氧酶(RuBisCO)小亚基RbcScDNA片段。方法:根据 莱茵衣藻、团藻、玉米等真核生物的RuBisCO小亚基RbcS氨基酸的高度保守序列ETRSYLPP和MNKLPMFG,设计 一对简并引物,采用RT PCR方法及该简并引物克隆杜氏盐藻RbcScDNA。PCR产物经T A克隆与T vector相连, 转化大肠杆菌JM109,随机挑取数个菌落,筛选鉴定,对阳性克隆进行测序分析。将测序结果推导成氨基酸序列进 行同源性比较。结果:得到的cDNA长度为209bp,编码69个氨基酸。推导的氨基酸序列与团藻、Chloromonassp. ANT3、衣藻、伞藻、菠菜、玉米的RbcS相比较,同源性分别为85%、84%、82%、76%、70%、69%。结论:所克隆的序 列为杜氏盐藻RbcScDNA片段。更多还原

【Abstract】 Aim: To obtain the small subunit (RbcS)cDNA fragment of ribulose-1,5-bisphosphate carboxylase/ oxygenase (RuBisCO) from Dunaliella salina. Methods: One pair of degenerate primer was designed according to conserved motifs of ETRSYLPP and MNKLPMFG of known RbcS and was used to amplify the RbcS cDNA fragment from the total RNA of Dunaliella salina. The resulting PCR product was cloned into pMD18-T vector and then transformed into E.coli JM109. Several clones were selected randomly and screened to determine their sequences. Homologous analysis of the deduced amino acid sequences was performed by BLAST and subsequently compared with GenBank data. Results: The obtained cDNA sequence was 209 base pairs in size, which encoded 69 amino acids. The sequence shared high identify with other RbcS Volvox carteri 85 %,Chloromonas sp. ANT3 84 %,Chlamydomonas reinhardtii 82 %,Acetabularia cliftonii 76 %,Spinacia oleracea 70 %,and Zea Mays 69 %, respectively. Conclusion: The cloned sequence is probably RbcS cDNA fragment from Dunaliella salina.更多还原