TY - JOUR
T1 - Isolation of RNA from bacterial samples of the human gastrointestinal tract
AU - Zoetendal, E.G.
AU - Booijink, C.C.G.M.
AU - Klaassens, E.S.
AU - Heilig, G.H.J.
AU - Kleerebezem, M.
AU - Smidt, H.
AU - de Vos, W.M.
PY - 2006
Y1 - 2006
N2 - The human gastrointestinal (GI) tract contains a complex microbial community that consists of numerous uncultured microbes. Therefore, nucleic-acid-based approaches have been introduced to study microbial diversity and activity, and these depend on the proper isolation of DNA, rRNA and mRNA. Here, we present an RNA isolation protocol that is suitable for a wide variety of GI tract samples. The procedure for isolating DNA from GI tract samples is described in another Nature Protocols article. One of the benefits of our RNA isolation protocol is that sampling can be performed outside the laboratory, which offers possibilities for implementation in large intervention studies. The RNA isolation is based on mechanical disruption, followed by isolation of nucleic acids using phenol:chloroform:isoamylalcohol extraction and removal of DNA. In our laboratory, this protocol has resulted in the isolation of rRNA and mRNA of sufficient quality and quantity for microbial diversity and activity studies. Depending on the number of samples, the sample type and the quenching procedure chosen, the whole procedure can be performed within 2.5¿4 h
AB - The human gastrointestinal (GI) tract contains a complex microbial community that consists of numerous uncultured microbes. Therefore, nucleic-acid-based approaches have been introduced to study microbial diversity and activity, and these depend on the proper isolation of DNA, rRNA and mRNA. Here, we present an RNA isolation protocol that is suitable for a wide variety of GI tract samples. The procedure for isolating DNA from GI tract samples is described in another Nature Protocols article. One of the benefits of our RNA isolation protocol is that sampling can be performed outside the laboratory, which offers possibilities for implementation in large intervention studies. The RNA isolation is based on mechanical disruption, followed by isolation of nucleic acids using phenol:chloroform:isoamylalcohol extraction and removal of DNA. In our laboratory, this protocol has resulted in the isolation of rRNA and mRNA of sufficient quality and quantity for microbial diversity and activity studies. Depending on the number of samples, the sample type and the quenching procedure chosen, the whole procedure can be performed within 2.5¿4 h
U2 - 10.1038/nprot.2006.143
DO - 10.1038/nprot.2006.143
M3 - Article
SN - 1754-2189
VL - 1
SP - 954
EP - 959
JO - Nature protocols
JF - Nature protocols
ER -