基于皱皮软海绵宏基因组的PKS基因筛选的研究  

PKS gene screening based on metagenome of Halichondria rugosa

作　　者：张戌升[1] 李志勇[1] 缪晓玲[1] 

机构地区：[1]上海交通大学生命科学技术学院海洋生物技术实验室,上海200240

出　　处：《微生物学报》2007年第3期526-528,共3页Acta Microbiologica Sinica

基　　金：国家"863计划"(2004AA628060)~~

摘　　要：提取皱皮软海绵及其共附生微生物的宏基因组总DNA,使用聚酮合酶(PKS)基因的酮酰合酶(KS)域引物PCR扩增PKS基因片段获得一条671bp的片段,以pUCm-T vector为载体将该基因片段克隆到大肠杆菌中,从阳性克隆中分离出PKS基因片段,测序推导出氨基酸序列。通过BLAST比对发现此氨基酸序列与红细菌目的Rhodobacterales bacterium PKS基因KS域的氨基酸序列有96%的同源性。通过基于氨基酸序列的系统发育分析,推测此筛选得到的PKS基因属于trans-AT型。本文首次证实了皱皮软海绵中存在细菌来源的PKS基因。Metagenome DNA was extracted from Halichondria rugosa which was collected from South China Sea and kept in -4℃. PKS gene fragment was amplified using PCR with KS domain primers in PKS gene. A DNA fragment about 671bp in length was obtained by PCR. The PCR product was measured by agrose gel electrophoresis. Then the product was recovered from gel and cloned into pUCm-T vector. After that vectors were transformed into competent cells （DHSa）. PKS gene fragment in positive clones was sequenced. Consequently, the corresponding amino acid sequence was deduced based on nucleotide sequence. BLAST analysis showed that the homology of this amino acid sequence with that deduced from KS domain of PKS gene in Rhodobacterales bacterium was up to 96 % . Phylogenetic analysis indicated that the obtained PKS gene belongs to trans-AT KS domains. Meanwhile the result demonstrated the diversity and differences of microorganisms associated with and around sponge in different sea area. It is the first time to find bacterial PKS gene in sponge Halichondria rugosa, which provide powerful proof to the microbial origin hypothesis of sponge active compounds. At the same time, this study lay basis for the utilization of uncultured microorganisms associated with sponge from the aspect of genes.

关 键 词：皱皮软海绵 宏基因组 聚酮合酶基因 PCR扩增 序列分析 

分 类 号：Q953[生物学—动物学]