[DNA extraction methods of compost for molecular ecology analysis].
Molecular ecology provides new techniques for studying compost microbes, and the DNA extraction is the basis of molecular techniques. Because of the contamination of humic acids, it turns to be more difficult for compost microbial DNA extraction. Three different approaches, named as lysozyme lysis, ultrasonic lysis and proteinase K lysis with CTAB, were used to extract the total DNA from compost. The detection performed on a nucleic acids and protein analyzer showed that all the three approaches produced high DNA yields. The agarose gel electrophoresis showed that the DNA fragments extracted from compost had a length of about 23 kb. A eubacterial 16S rRNA gene targeted primer pair (27F and 1 495R) was used for PCR amplification, and all the samples got almost the full length 16S rDNA sequence (about 1.5 kb). After digested by restriction endonucleases (Hae Ill and Alu I), the restriction map showed relatively identical microbial diversity in the DNA, which was extracted by the three different approaches. All the compost microbial DNA extracted by the three different approaches could be used for molecular ecological study, and researchers should choose the right approach for extracting microbial DNA from compost based on the facts.
Yang
Zhao-Hui
ZH
College of Environmental Science and Engineering, Hunan University, Changsha 410082, China. yzh@hnu.cn
Xiao
Yong
Y
Zeng
Guang-Ming
GM
Liu
Yun-Guo
YG
Deng
Jiu-Hua
JH
D001419:Bacteria / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics / Q000302:isolation & purification*; D004463:Ecology / Q000379:methods; D004587:Electrophoresis, Agar Gel; D005285:Fermentation; D008967:Molecular Biology / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012037:Refuse Disposal; D012722:Sewage / Q000382:microbiology*
D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S; D012722:Sewage