潮间带污泥中厌氧发酵产氢混合菌群组成与Fe-氢酶基因多样性分析  

DIVERSITY ANALYSIS OF MICROBIAL COMMUNITY AND Fe-HYDROGENASE GENES OF FERMENTATIVE MIXED CULTURE FROM INTERTIDAL SLUDGE

作　　者：刘洪艳[1,2] 王广策[2] 朱大玲[1] 潘光华[1] 

机构地区：[1]天津科技大学海洋科学与工程学院天津市海洋资源与化学重点实验室,天津300457 [2]中国科学院海洋研究所,青岛266071

出　　处：《海洋与湖沼》2012年第1期180-184,共5页Oceanologia Et Limnologia Sinica

基　　金：国家自然科学基金项目,40906074号;天津市自然科学青年基金项目,12JCQNJC04200号;天津科技大学科学研究基金资助项目,20110101号

摘　　要：以采自天津海水浴场潮间带的污泥为研究材料,将污泥热休克处理后富集混合菌群进行厌氧发酵产氢,测定混合菌群在发酵过程中累积产氢量和相关产氢指标(吸光度、酸碱度和氧化还原电位)的变化。结果表明,热休克处理后富集的产氢混合菌群产氢量为0.41mol H2/mol葡萄糖。产氢结束后,利用变性梯度凝胶电泳(DGGE)分析混合菌群组成和Fe-氢酶基因的多样性。混合菌群基因组DNA的16S rRNAV3区扩增产物经过DGGE电泳分离,结果得到6条丰度比较高的条带,测序结果表明优势菌为Clostridium sp.。利用梭菌氢酶基因保守性引物克隆混合产氢菌群中梭菌Fe-氢酶基因,扩增产物经DGGE电泳分离,电泳结果表明混合菌群中Fe-氢酶基因分为3个分类单元,NCBI-BLAST比对结果与Clostridium roseum和Clostridium perfringens的Fe-氢酶基因相似度分别为79%和98%。分析产氢混合菌群组成和Fe-氢酶基因多样性,可以为扩大产氢微生物种质资源奠定基础。Sludge sample, collected from intertidal zone of a bathing beach in Tianjin, was pretreated by heat-shock method for enrichment of hydrogen-producing microbial community. The hydrogen production and indicators of hydrogen production （OD600, pH and ORP values） were measured. The results showed that the efficiency of hydrogen production of the heat-shock pretreated sludge was 0.41mol H2/mol glucose. At the end of fermentation, 16S rRNA gene and Fe-hydrogenase gene of the mixed culture were PCR amplified and analyzed by DGGE （denaturing gradient gel electrophoresis） respectively. For 16S rRNA gene six special bands were obtained. The sequencing results showed that the dominate bacterium of the mixed culture belong to Clostridium sp.. Heat-shock pretreatment was favorable to enrich the most known hydrogen-producing bacterium, i.e. Clostridium sp.. As for the Fe-hydrogenase gene of Clostridium three opera- tional taxonomical units （OUTs） were obtained. The segments of Fe-hydrogenase gene were cloned and sequenced. NCBI blast result indicates that the segments of Fe-hydrogenase gene were 79% and 98% identical to that of Clostridium roseum and Clostridium perfringens respectively. The analysis about composition of microbial community and diversity of Fe-hydrogenase gene contribute to enlarge the resource of hydrogen-producing bacteria group.

关 键 词：混合菌群 产氢量 16S RRNA Fe-氢酶基因 变性梯度凝胶电泳 

分 类 号：Q933[生物学—微生物学]