Comparison of methods for total community DNA extraction and purification from compost.
The differences on DNA yield and purity of three different DNA extraction protocols were compared with regard to the use for PCR and other molecular analyses. Total DNA was extracted from compost by the three protocols, and then was purified by spin-bind cartridges after being precipitated by PEG8000. The detection performed on a nucleic acid and protein analyzer showed that all three methods produced high DNA yields. The agarose gel electrophoresis showed that the fragments of crude and purified DNA had a length of about 23 kb. A eubacterial 16S rRNA gene-targeted primer pair was used for PCR amplification, and full length 16S rDNAs were amplified from all the purified DNA samples. After being digested by restriction endonucleases, the restriction map of amplified rDNA showed identical genetic diversity. The products of PCR using primer pair GC341F and 907R were also used for denaturing gradient gel electrophoresis analysis. The results indicated that high-quality DNA was extracted from compost by the three protocols, and each of the protocols is adapted to extract microbial genome DNA from compost expediently and cheaply.
Yang
Zh H
ZhH
College of Environmental Science and Engineering, Hunan University, Changsha, Hunan Province, China. yzh@hnu.cn
Xiao
Y
Y
Zeng
G M
GM
Xu
Zh Y
ZhY
Liu
Y Sh
YSh
D004247:DNA / Q000302:isolation & purification*; D016172:DNA Fingerprinting; D004275:DNA, Ribosomal / Q000235:genetics / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D008967:Molecular Biology / Q000379:methods*; D009691:Nucleic Acid Denaturation; D012150:Polymorphism, Restriction Fragment Length; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012987:Soil; D012988:Soil Microbiology
D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S; D012987:Soil; D004247:DNA