限制性片段长度多态性分析(ARDRA)方法对重金属污染土壤中细菌群落多样性的研究

Use of amplified ribosomal DNA restriction analysis to study microbial diversity in soils impacted by heavy metals

限制性片段长度多态性分析(ARDRA)是用限制性内切酶消化扩增后的细菌16SrDNA,通过凝胶电泳分析微生物种群多样性的分子标记技术,该技术在微生物生态学领域有着广泛的应用.实验采用2种途径评价了这种方法应用于研究土壤中细菌种群多样性的可行性.通过对未污染土壤细菌16SrDNA的克隆,获得了经内切酶HaeⅢ消化的不同16SrDNA的ARDRA类型,并对测序的19个克隆子建立了系统发育树.此外,还对不同Cd和Pb污染水平的土壤微生物进行了群落水平上的ARDRA分析.结果表明,5mg·kg-1Cd和500mg·kg-1Pb污染对微生物群落组成有一定影响,但此范围内的重金属含量对微生物群落多样性没有相关性影响.

Amplified ribosomal DNA restriction analysis （ARDRA） is a DNA fingerprinting technique based on restriction endonuclease digestion of the amplified 16S ribosomal DNA, followed by agarose gel electrophoresis. In this study, the suitability of this method to investigate the bacterial communities in soils was evaluated. ARDRA patterns using enzyme digestion （HaeⅢ） of cloned 16S RNA gene sequences from uncontaminated soil were obtained, and a phylogenetic tree was built according to the 19 sequenced clones. In addition, ARDRA on the whole community level was applied to soils contaminated with different amounts of Cd and Pb. The results indicated that 5 mg·kg^-1 Cd and 500 mg·kg^-1 Pb affected soil microbial community structure, however, no relationship, a positive relationship was observed between heavy metal concentration and microbial diversity, and the largest diversity index appeared in the most heavily polluted soils.