厌氧颗粒污泥PCR-DGGE图谱的优化研究

The Optimization of PCR-DGGE Map of Anaerobic Granular Sludge

旨在为厌氧颗粒污泥微生物多样性研究的开展提供指导，实验以处理果汁废水的厌氧颗粒污泥为研究对象，对PCR—DGGE实验条件进行了优化研究。结果表明：1）采用巢式PCR对厌氧颗粒污泥内细菌与古菌进行扩增，古菌第二轮扩增时退火温度58℃，延伸时间408得到的PCR产物可满足DGGE的要求；2）古菌与细菌的DGGE最优变性剂梯度分别为40％-60％和30％～45％，电泳条件均为80V，16h；3）改进的强碱硝酸银染色法为高效低污染的染色方法。

The optimal conditions of PCR-DGGE technique was studied in this paper with anaerobic granules from fruit juice wastewater treatment as object. The results indicated that： 1. the DNA of archaca and bacteria from the granules was amplified using nest-PCR, for the second round amplification, the PCR products of archaca could meet the requirement of DGGE when annealing temperature was 58 ℃, and extension time 40 s; 2. the optimal denaturant gradient of DGCE for archaea and bacteria were 40% - 60% and 30% - 45% respectively, while the best electrophoretic conditions for both of the archaea and bacteria were 80V for 16 h; 3. the modified strong alkali silver nitrate staining protocol was the effetive stai- ning method with low pollution.