新的J超家族芋螺毒素的克隆与进化分析

摘要

Objective: To identify and analyze new J-superfamily conotoxins from six Conus species from the South China Sea. Methods: Total RNA was extracted from the venom duct of Conus species and was reverse transcribed into cDNA. The sequences of J-superfamily conotoxins were cloned by 3'-rapid amplification of cDNA ends(RACE) and nest PCR method. The expected bands were then connected into the pMD18-T vector and transformed into E.coli DH5α competent cells. Applying the NCBI database, novel J-superfamily conotoxins were identified. The characterization and phylogenetic tree of novel J-superfamily conotoxins were analyzed using BioEdit, ClustalW and Mega5.05 software. Results: Twelve novel J-superfamily conotoxins with different amino acids residues were identified from six Conus species. The analysis of phylogenetic tree showed that these novel toxins located in the different branch. Conclusion: The novel J-superfamily conotoxins exhibit less homology to the reported, and belong to the new members of J-superfamily conotoxi%目的:从中国南海芋螺中克隆新的J超家族芋螺毒素,并进行序列和进化分析.方法:以芋螺毒腺管总RNA为模板,采用3'-RACE及巢式PCR的方法扩增J超家族芋螺毒素基因,并将得到的目的基因与pMD18-T载体连接并转化大肠杆菌DH5α,经测序比对后,获得新的J超家族毒素,利用软件BioEdit、ClustalX及Mega5.05进行进化分析.结果:获得12个新的J超家族芋螺毒素前体肽序列,其氨基酸组成具有新颖性,在进化树上与已报道的J超家族毒素处于不同的进化分支.结论:12个新的J超家族毒素与已报道的毒素序列之间的同源性较低,是J超家族新成员.