ABSTRACT
How to get functional gene from uncultured-microbiology is the hotspot content of microbial ecology. What the most important is how to obtain the pure and integrated genomic DNA. An efficient, nonselective extraction method to gain chromosomal DNA from eight kinds of bacteria was introduced. Amount DNA released by hot-detergent gave the highest DNA yields from different G + and G- bacteria. Running 20 hours by PFGE mode, the size of total DNA is over 23kb. The pure DNA could be digested by Hind Ⅲ and used in PCR. The total environmental DNA also can be extracted from soil by the same method. As a result it showed a new way for the environmental DNA extraction.
ABSTRACT
Methods for studying the population diversity of microorganism in activated sludge usually require enrichment of bacterial genome.The efficient information on microbial species composition provided and shifted in diversity revealed are dependent on the effective DNA recovery technique.The method was based on washing by alkaline phosphate buffer and digestion with extended heating of the activated sludge suspension in the presence of lysozyme and freeze-thawing in high-salt-SDS buffer.The extraction was tested for four activated sludge differing in places and dates.The DNA fragment from all sludge was integrity.DNA yields ranged from 105 to 823 ?g/g sludge and were of sufficient purity for PCR-based 16S ribosomal DNA analysis and restriction digested.In general,all methods produced DNA pure were not enough for PCR amplification and libraries construction.As basis of experimental goals,the study provides an appropriate extraction method of microbial DNA in sludge.