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作 者 ：高平平,赵立平

期 刊 ：生态学报 2002年 22卷 11期 页码：2015~2019

关键词：活性污泥；DNA提取；PCR扩增；

Keywords:activated sludge, DNA extraction, PCR amplification,

摘 要 ：活性污泥样品经液氮速冻、沸水浴融化、溶菌酶处理和 SDS裂解后 ,99%以上细胞裂解。所提取的 DNA经琼脂糖凝胶电泳检测和荧光法浓度测定 ,其片断大小在 2 0 kb左右 ,产量可达 1 .75 6± 0 .1 mg/g MLSS。样品 ABS2 6 0 nm/ABS2 80 nm的比值为 1 .96± 0 .2。以提取的总 DNA为模板 ,进行细菌核糖体小亚基 1 6Sr DNA基因 V3区和多组分苯酚羟化酶大亚基基因 (Lm PHs)的 PCR扩增 ,均获得成功 ,为活性污泥中微生物群落的分子生态学研究提供了一种简便、可靠的 DNA提取方法。

Abstract:A rapid and less instrument\|dependent protocol for direct extraction of DNA from activated sludge sample was developed. To optimize the cell lysis step, four cell lysis treatments (Freeze\|thaw, lysozyme, SDS , and thermal shock +lysozyme + SDS) were used to disrupt dispersed cells and release DNA. The efficiency of cell lysis was assessed microscopically after each treatment step. The quality and quantity of extracted DNA was determined by using both agarose gel electrophoresis and spectrophotometric meas...

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