杜氏盐藻泛素基因cDNA的克隆及系统进化分析  

Cloning and phylogenetic analysis of ubiquitin cDNA from Dunaliella salina

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作  者:刘玲玲[1] 李杰[1] 宋贤瑞[1] 梁建阳[1] 薛乐勋[1] 

机构地区:[1]郑州大学细胞生物学研究室,郑州大学第一附属医院,郑州450052

出  处:《生物学杂志》2008年第2期19-22,共4页Journal of Biology

基  金:国家自然科学基金资助项目(30270031,30300208)

摘  要:根据玉米,水稻等物种泛素序列设计一对简并引物。提取杜氏盐藻细胞的总RNA,利用RT-PCR方法扩增盐藻泛素基因的cDNA片断,回收两个长度不同的片断ubi-1和ubi-2,将其克隆到pMD18-T载体上,测序后进行序列分析,为克隆杜氏盐藻泛素基因的cDNA序列并进行进化分析。结果ubi-1经测序后得到一个完整拷贝(228bp)和一个不完整的泛素cDNA序列(191bp)。Ubi-2经测序后得到两个拷贝(556bp)和一个不完整的泛素cDNA序列(191bp)。盐藻3个不同拷贝泛素cDNA序列之间存在差异,但所编码氨基酸序列相同。盐藻泛素cDNA序列与其他物种的泛素cDNA序列具有高的同源(70%~85%),所推导的氨基酸序列与其他物种仅存在1~2个氨基酸的差异。进化分析显示,所分离的盐藻泛素基因与两个模式生物果蝇和衣藻的泛素基因共处一个进化支.彼此亲缘关系最近。盐藻泛素基因与其他物种的泛素基因可能来自共同的“祖先”基因,在进化中高度保守。To clone and analyze cDNA sequences of the ubiquitin gene derived from Dunaliella salina (D. salina), degenerate primers were designed based on conserved amino acid sequences of maize, flee. RT-PCR was performed to amplify eDNA fragments of ubiquitin gene from D. salina. The amplified eDNA fragments named ubi-1 and ubi-2 were cloned into the vector pMD18-T and sequenced. Results showed that the eDNA fragment named ubi-1 contains a 228 bp ubiquitin unit and a 191 bp partial eDNA fragment, eDNA fragment ubi-2 contains two 228 bp ubiquitin units and a 191 bp partial eDNA fragment. Sequence analysis of the cloned three ubiquitin units showed variation at the nucleotide level, but no variation in the encoded amino acid sequences. Sequences alignment analysis showed that the ubiquitin eDNA unit from D. salina shared high homology with that from other species (70%-85% ), and there were only 1-2 encoded amino acid sequences difference between that from D. salina and other species. The analysis of evolution relationship showed that the ubiquitin gene from D. salina was more closely related to that from model organisms D. melanogaster and C. reinhardtii and was highly conserved during evolution.

关 键 词:杜氏盐藻 泛素基因 进化分析 

分 类 号:Q754[生物学—分子生物学]

 

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