新的J超家族芋螺毒素的克隆与进化分析

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。

Cloning and Phylogenetic Analysis of Novel J-Superfamily Conotoxins

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 tran scribed 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 trans formed into E.coli DH5α competent cells.Applying the NCBI database,novel J-superfamily conotoxins were identi fied.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 resi dues were identified from six Conus species.The analysis of phylogenetic tree showed that these novel toxins locat ed 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 conotoxins.