PMID- 20505753
OWN - NLM
STAT- MEDLINE
DCOM- 20110104
LR  - 20100921
IS  - 1751-7370 (Electronic)
IS  - 1751-7362 (Linking)
VI  - 4
IP  - 10
DP  - 2010 Oct
TI  - Fixation-free fluorescence in situ hybridization for targeted enrichment of 
      microbial populations.
PG  - 1352-6
LID - 10.1038/ismej.2010.73 [doi]
AB  - We modified the standard ribosomal RNA-targeted fluorescence in situ 
      hybridization (FISH) protocol by removing the fixation steps to allow recovery of 
      unmodified nucleic acids. Using this method, hybridized cells could be visualized 
      in two bioreactor sludges and termite hindgut samples by epifluorescence 
      microscopy. We then targeted one bacterial and one archaeal population in the 
      sludge samples with group-specific oligonucleotide probes using in-solution 
      fixation-free FISH and sorted hybridized populations using fluorescence-activated 
      cell sorting (FACS). We could show that sorted populations were highly enriched 
      for the target organisms based on 16S rRNA gene sequencing, thus confirming probe 
      specificity using the modified FISH protocol. This approach should facilitate 
      subsequent genomic sequencing and analysis of targeted populations as DNA is not 
      compromised by crosslinking during fixation.
FAU - Yilmaz, Suzan
AU  - Yilmaz S
AD  - Microbial Ecology Program, DOE Joint Genome Institute, Walnut Creek, CA, USA.
FAU - Haroon, Mohamed F
AU  - Haroon MF
FAU - Rabkin, Brian A
AU  - Rabkin BA
FAU - Tyson, Gene W
AU  - Tyson GW
FAU - Hugenholtz, Philip
AU  - Hugenholtz P
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, Non-P.H.S.
DEP - 20100527
PL  - England
TA  - ISME J
JT  - The ISME journal
JID - 101301086
RN  - 0 (RNA, Archaeal)
RN  - 0 (RNA, Bacterial)
RN  - 0 (RNA, Ribosomal, 16S)
RN  - 0 (Sewage)
SB  - IM
MH  - Animals
MH  - Archaea/genetics/*isolation & purification
MH  - Bacteria/genetics/*isolation & purification
MH  - Bioreactors/microbiology
MH  - Flow Cytometry/methods
MH  - Gastrointestinal Tract/microbiology
MH  - In Situ Hybridization, Fluorescence/*methods
MH  - Isoptera/microbiology
MH  - Microbiological Techniques/*methods
MH  - Microscopy, Fluorescence/*methods
MH  - RNA, Archaeal/genetics
MH  - RNA, Bacterial/genetics
MH  - RNA, Ribosomal, 16S/genetics
MH  - Sewage/microbiology
EDAT- 2010/05/28 06:00
MHDA- 2011/01/05 06:00
CRDT- 2010/05/28 06:00
PHST- 2010/05/28 06:00 [entrez]
PHST- 2010/05/28 06:00 [pubmed]
PHST- 2011/01/05 06:00 [medline]
AID - ismej201073 [pii]
AID - 10.1038/ismej.2010.73 [doi]
PST - ppublish
SO  - ISME J. 2010 Oct;4(10):1352-6. doi: 10.1038/ismej.2010.73. Epub 2010 May 27.