| 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 rRNA V3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。 |
| 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 |
| DOI：10.11693/hyhz201201029029 |
| 分类号: |
| 基金项目: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。 |
|
| DIVERSITY ANALYSIS OF MICROBIAL COMMUNITY AND Fe-HYDROGENASE GENES OF FERMENTATIVE MIXED CULTURE FROM INTERTIDAL SLUDGE |
|
LIU Hong-Yan1,2, WANG Guang-Ce2, ZHU Da-Ling1, PAN Guang-Hua1
|
|
1.College of Marine Science and Engineering, Tianjin University of Science and Technology, Tianjin Key Laboratory of Marine Resources and Chemistry;2.Institute of Oceanology, Chinese Academy of Sciences
|
| Abstract: |
| 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 operational 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. |
| Key words: Mixed culture, Hydrogen production, 16S rRNA, Fe-hydrogenase gene, Denatured gradient gel electrophoresis |
