An effective method for isolation of DNA from pig faeces and comparison of five different methods.
Polymerase chain reaction (PCR) detection of microorganism in faecal specimens is hampered by poor recovery of DNA and by the presence of PCR inhibitors. In this paper, we describe a new modified method for extracting PCR-quality microbial community DNA from pig faecal samples, which combines the pretreatment with polyformaldehyde, and subsequent DNA lysis in the presence of CTAB, salt, PVP, and beta-mercaptoethanol, followed by isolation of nucleic acids using chloroform (no phenol) based protocol. The method resulted in a 1.3- to 11-fold increase in DNA yield when compared to four other widely used methods. Genomic DNA extracted from all five methods was assessed by both agarose gel electrophoresis and polymerase chain reaction for amplification of 16S rDNA specific fragments. The results showed that the improved method represented a reproducible, simple, and rapid technique for routine DNA extraction from faecal specimens and was notably better than using the QIAamp DNA Stool Mini Kit.
Tang
Jun-ni
JN
Bioengineering Research Center for Animal Disease Prevention and Control, School of Life Science, Sichuan University, Chengdu, China.
Zeng
Zhi-guang
ZG
Wang
Hong-ning
HN
Yang
Tai
T
Zhang
Peng-ju
PJ
Li
Yu-ling
YL
Zhang
An-yun
AY
Fan
Wen-qiao
WQ
Zhang
Yi
Y
Yang
Xin
X
Zhao
Su-jun
SJ
Tian
Guo-bao
GB
Zou
Li-kou
LK
D000818:Animals; D001419:Bacteria / Q000235:genetics / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification*; D004275:DNA, Ribosomal / Q000235:genetics; D005243:Feces / Q000382:microbiology*; D005821:Genetic Techniques; D016133:Polymerase Chain Reaction; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D013552:Swine
D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S