PMID: 9546190Apr 18, 1998Paper

Development of a direct in situ PCR method for detection of specific bacteria in natural environments

Applied and Environmental Microbiology
K TaniM Nasu

Abstract

We applied HNPP (2-hydroxy-3-naphthoic acid-2'-phenylanilide phosphate) to direct in situ PCR for the routine detection of specific bacterial cells at the single-cell level. PCR was performed on glass slides with digoxigenin-labeled dUTP. The digoxigenin-labeled PCR products were detected with alkaline phosphatase-labeled antidigoxigenin antibody and HNPP which was combined with Fast Red TR. A bright red fluorescent signal was produced from conversion to HNP (dephosphorylated form) by alkaline phosphatase. We used the ECOL DNA primer set for amplification of ribosomal DNA of Escherichia coli to identify cells specifically at the single-cell level in a bacterial mixture. High-contrast images were obtained under an epifluorescence microscope with in situ PCR. By image analysis, E. coli cells in polluted river water also were detected.

References

Apr 25, 1991·Nucleic Acids Research·G J OlsenC R Woese
Oct 5, 1990·Journal of Molecular Biology·S F AltschulD J Lipman
Apr 1, 1988·Proceedings of the National Academy of Sciences of the United States of America·W R Pearson, D J Lipman
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Citations

Oct 27, 2001·Applied and Environmental Microbiology·T HoshinoY Inamori
Feb 10, 2004·Applied and Environmental Microbiology·Michinari SunamuraAkihiko Maruyama
Sep 10, 2005·Applied and Environmental Microbiology·Takehiko KenzakaMasao Nasu
Oct 1, 2010·Critical Reviews in Microbiology·Jacques TheronMichele de Kwaadsteniet
Jan 5, 2002·Applied and Environmental Microbiology·Katsuji TaniMasao Nasu
Sep 9, 2004·Microbiology and Molecular Biology Reviews : MMBR·Byron F Brehm-Stecher, Eric A Johnson
Nov 5, 2005·Applied and Environmental Microbiology·Takehiko KenzakaMasao Nasu
Sep 1, 2005·Critical Reviews in Food Science and Nutrition·Ioannis S Arvanitoyannis, Maria Van Houwelingen-Koukaliaroglou
Dec 8, 2005·Applied and Environmental Microbiology·Fumito MaruyamaMasao Nasu
Mar 27, 2007·Applied and Environmental Microbiology·Takehiko KenzakaMasao Nasu
Mar 16, 2007·Clinical and Vaccine Immunology : CVI·C Maldonado GaldeanoG Perdigón
Sep 5, 2008·Current Protocols in Cytometry·Jonathan Porter
Sep 24, 2008·Journal of Medical Microbiology·Huili HuYonghong Yang
May 29, 1999·Letters in Applied Microbiology·K KurokawaM Nasu
Feb 14, 2003·Biotechnology and Bioengineering·Ramaiah Sachidanandham, Karina Yew-Hoong Gin
Aug 7, 2003·Applied and Environmental Microbiology·Fumito MaruyamaMasao Nasu
Nov 19, 2019·Frontiers in Microbiology·Qin MaSenthil Subramanian
Sep 23, 2000·Journal of Microbiological Methods·J Porter, R W Pickup
Jan 5, 2002·Journal of Microbiological Methods·Annie RompréPatrick Laurent
Oct 16, 1999·Journal of Microbiological Methods·A Vaid, A H Bishop
Dec 1, 1999·Journal of Microbiological Methods·V L WilsonP LaRock

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