垃圾填埋场中真细菌群落16S rRNA基因的分析

回灌式垃圾填埋场渗滤液中真细菌群落的多样性同样通过不依赖于微生物培养的分析而获得。利用特异性的引物对,选择性地扩增渗滤液DNA中的真细菌16S rRNA基因片断(16S rDNA),并用于构建16S rDNA克隆文库。文库内真细菌16S rDNA的遗传多样性通过限制片断长度多态性分析(RFLP,限制性内切酶Hin PII和Msp I)而获得。初步的结果表明,随机选出的200个真细菌克隆子被分为147个不同的RFLP型(组),当中丰度最高的两个型均仅含有9个克隆子,两者共占所有被分析克隆子的不到10%;克隆子数≥2的型共有21个(包括以上2个丰度最高的型),它们共代表74个克隆子,占所有被分析克隆子的37%;而剩下的126个型均只含有1个克隆子,它们共占整个基因文库的63%。由此可见,李坑垃圾渗滤液中的真细菌具有非常复杂的群落结构。

Characterization of Eubacteria Community in Landfill by Determining Restriction Fragment Length Polymorphisms of 16S rRNA Genes

By using a culture-independent molecular approach,we analyzed the community structure of Bacteria in the effluent leachate of a full-scale recirculating landfill.Bacterial 16S rRNA genes(rDNA) were amplified directly from leachate DNA by PCR with universally conserved and Bacteria-specific rDNA primers and cloned.Genetic variation of bacterial 16S rDNAs in the clone library was gained by a restriction fragment length polymorphism analysis(RFLP).The cloned 16S rDNA fragments were reamplified,and restriction analysis was performed following separate digestion with enzymes Hin PII and Msp I.A total of 200 randomly selected recombinant clones were examined,and they were finally clustered into 147 unique groups(RFLP patterns).The two most frequent groups,both including nine clones,accounted for less than 10% of the total bacterial clones analyzed;the 21 more frequent groups,each including ≥2 clones,represented 74 clones and accounted for 37% of the total bacterial clones;and each of the remaining 126 groups was represented by a single clone.These preliminary results showed that the community structure of bacteria in the landfill leachate was far more complex and variable than expected and thus could be challenging to control.