IS-pro: high-throughput molecular fingerprinting of the intestinal microbiota.
The human intestinal microbiota is known to play an important role in human health and disease, and with the advent of novel molecular techniques, disease-specific variations in its composition have been found. However, analysis of the intestinal microbiota has not yet been applicable in large-scale clinical research or routine diagnostics because of the complex and expensive nature of the techniques needed. Here, we describe a new PCR-based profiling technique for high-throughput analysis of the human intestinal microbiota, which we have termed IS-pro. This technique combines bacterial species differentiation by the length of the 16S-23S rDNA interspace region with instant taxonomic classification by phylum-specific fluorescent labeling of PCR primers. We validated IS-pro in silico, in vitro, and in vivo, on human colonic biopsies and feces, and introduced a standardized protocol for data analysis. IS-pro is easy to implement in general clinical microbiological laboratories with access to capillary gel electrophoresis, and the high-throughput nature of the test makes analysis of large numbers of samples feasible. This combination renders IS-pro ideally suited for use in clinical research and routine diagnostics.
Budding
A E
AE
Department of Medical Microbiology and Infection Control, VU University Medical Center, Amsterdam, The Netherlands. d.budding@vumc.nl
Grasman
M E
ME
Lin
F
F
Bogaards
J A
JA
Soeltan-Kaersenhout
D J
DJ
Vandenbroucke-Grauls
C M J E
CM
van Bodegraven
A A
AA
Savelkoul
P H M
PH
D001419:Bacteria / Q000145:classification / Q000235:genetics*; D044822:Biodiversity; D016172:DNA Fingerprinting / Q000379:methods*; D021903:DNA, Ribosomal Spacer / Q000235:genetics; D019075:Electrophoresis, Capillary / Q000379:methods; D005243:Feces / Q000382:microbiology*; D006801:Humans; D007422:Intestines / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods; D015203:Reproducibility of Results; D013045:Species Specificity
D021903:DNA, Ribosomal Spacer