Effect of PCR template concentration on the composition and distribution of total community 16S rDNA clone libraries

Molecular Ecology
D P ChandlerF J Brockman

Abstract

Total DNA from sediment samples was isolated by a direct lysis technique. Purified DNA was used as template either undiluted or diluted 1:10 prior to polymerase chain reaction (PCR) amplification of 16S rRNA genes. Full-length inserts were analysed for restriction fragment length polymorphisms (RFLP) with the enzyme Cfo1, and the resulting distribution and abundance of RFLP patterns compared between the undiluted and diluted PCR reactions. Results indicate that for low PCR template concentrations, in the range from a few picograms to tens of picograms DNA, proportional representation of specific RFLP types was not reproducible upon template dilution, confirming that PCR amplification of 16S rDNA cannot be used directly to infer microbial abundance. In particular, only 15-24% of the RFLP types recovered from a sample were present in both the undiluted and diluted extracts. We propose that very low template concentrations in the PCR generate random fluctuations in priming efficiency, which led to the contrast in RFLP types observed in the libraries from the undiluted and diluted extracts.

Citations

Apr 6, 2006·Microbial Ecology·Jacques J SmithDonald A Cowan
Mar 9, 2007·Microbial Ecology·Andreas NockerAnne K Camper
Sep 15, 2010·Microbial Ecology·Anita PokharelDittmar Hahn
Aug 24, 2010·Applied Microbiology and Biotechnology·Antti Juhani RissanenMarja Tiirola
Nov 8, 2011·Journal of Microbiological Methods·Katja EngelJosh D Neufeld
May 30, 2012·Journal of Microbiological Methods·Paul M RueggerJames Borneman
Jun 19, 2001·FEMS Microbiology Letters·G C BakerA R Suharto
Jun 11, 1999·Current Opinion in Biotechnology·A G O'Donnell, H E Gorres
Oct 20, 2005·Journal of Bioscience and Bioengineering·K Watanabe, P W Baker
Oct 20, 2005·Journal of Bioscience and Bioengineering·Takahiro Kanagawa
Mar 19, 2002·Wound Repair and Regeneration : Official Publication of the Wound Healing Society [and] the European Tissue Repair Society·C E DaviesD W Thomas
Dec 11, 2007·Applied and Environmental Microbiology·Valeria A TorokRobert J Hughes
Mar 4, 2008·Applied and Environmental Microbiology·Sarah R Carrino-Kyker, Andrew K Swanson
Apr 14, 2009·Applied and Environmental Microbiology·Oleg PaliySonia Michail
Aug 5, 2000·Applied and Environmental Microbiology·D P ChandlerF J Brockman
Jul 31, 2002·Applied and Environmental Microbiology·David W ReedFrederick S Colwell
Nov 7, 2003·Applied and Environmental Microbiology·Jiangrang LuMargie D Lee
Dec 3, 2004·Applied and Environmental Microbiology·Shinya KurataYoichi Kamagata
Mar 10, 2004·Applied and Environmental Microbiology·Grieg F StewardJonathan P Zehr
Mar 28, 2013·BMC Ecology·Wouter F D van DongenRichard H Wagner
Nov 15, 2011·Microbes and Environments·Chiachi HwangWen-Tso Liu
Jul 11, 2013·World Journal of Microbiology & Biotechnology·N WatanarojanapornN Teaumroong
Dec 10, 1999·Proceedings of the National Academy of Sciences of the United States of America·I KroesD A Relman

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