海洋放线菌(Salinispora arenicola)基因转移系统的建立

Construction of the gene transfer system of marine-derived actinomycetes Slinispora arenicola

该实验旨在建立海洋放线菌Salinispora arenicola基因转移系统,以便基因敲除和外源基因表达等遗传操作。以整合型质粒pSET152和p IB139为出发质粒,通过接合转移构建了海洋放线菌Salinispora arenicola的基因转移系统。结果表明,25μg/m L阿泊拉霉素可有效筛选接合子,大肠杆菌与孢子的比例为8:1时可获得较多的转化子。经聚合酶链式反应(PCR)验证,质粒成功整合到菌株海洋放线菌S.arenicola基因组中,接合子经多次传代后,导入的质粒pSET152和pIB139仍稳定整合于接合子基因组上。成功构建了海洋放线菌S.arenicola接合转移系统。

The objective of this study is to estabiish the gene transfer system of marine-derived actinomycetes Saiinispora arenicoia,which can be used for genetic manipuiations such as gene knock-out and expression of foreign genes.Using integrated piasmid pSET152 with pIB139 as originai piasmid,the gene transfer system of actinomycetes S.arenicoia was constructed by conjugating transfer.Resuits showed that 25^g/mi apramycin may be used to efficientiy screening conjugants,and more transformant can be acquired with Escherichia coii to spores ratio 8:1.PCR verification reveaied that exogenous piasmid was successfuiiy integrated in the chromosomai DNA of actinomycetes S.arenicoia.After muitipie generations of zygotes,the transformed piasmid pSET152 and pIB139 of conjugants were stabiy integrated in the zygote genome.The S.arenicoia conjugationai transfer system was successfuiiy constructed.