African Journal of
Microbiology Research

  • Abbreviation: Afr. J. Microbiol. Res.
  • Language: English
  • ISSN: 1996-0808
  • DOI: 10.5897/AJMR
  • Start Year: 2007
  • Published Articles: 5233

Full Length Research Paper

Impact of different polymerase chain reaction (PCR) strategies on denaturing gradient gel electrophoresis-based analysis of bacterial communities in soils/sediments from the Northern Jiangsu Oil Field, China

Xiao-Ying Wu1,2, Xiao-Ming Xu2, Shao-Ying Fu3, Jian-Ping Yuan4, Juan Peng4, Teng-Peng Yan2, Chou-Fei Wu4* and Jiang-Hai Wang2*
1School of Life Sciences, Sun Yat-Sen University, Guangzhou 510275, People’s Republic of China. 2Guangdong Province Education Department Key Laboratory of Marine Petroleum Exploration and Development, School of Marine Sciences, Sun Yat-Sen University, Guangzhou 510006, People’s Republic of China. 3Guangzhou Marine Geological Survey, Guangzhou 510760, People’s Republic of China. 4Guangdong Provincial Key Laboratory of Marine Resources and Coastal Engineering, School of Marine Sciences, Sun Yat-Sen University, Guangzhou 510006, People’s Republic of China.
Email: [email protected], [email protected]

  •  Accepted: 05 October 2012
  •  Published: 13 November 2012

Abstract

The selection of polymerase chain reaction (PCR) strategies is vital to a successful assessment of bacterial communities in soils/sediments by 16S rRNA gene-based denaturing gradient gel electrophoresis (DGGE) analysis. To obtain reliable information of the bacteria communities in soils/sediments from the Northern Jiangsu Oil Field (NJOF), the impact of six PCR strategies on DGGE analysis has been investigated. The results showed that one-step PCR approach with primer set 341f⊥GC/534r (Strategies 1 and 2) was not suitable for 16S rRNA gene amplification of the bacterial communities in the NJOF soils/sediments before DGGE analysis due to its non-specific DNA amplification and low efficiency of 16S rRNA gene amplification. Strategy 6 (one nested PCR approach with primer sets 27f/907r and 341f⊥GC/534r with a purification procedure) could be the most accurate assessment of community diversities, but only be suitable to perform DGGE analysis for a few samples and not for high-throughput DGGE analysis because it was time-consuming; and Strategies 5 (one nested PCR approach with a dilution procedure) and 3 (one two-step PCR approach with primer sets 341f /534r and 341f⊥GC/534r) could provide similar information on the bacterial diversity of the NJOF soils/sediments without a purification procedure comparing with Strategy 6. Therefore, we prefer to recommend Strategies 5 and 3 for high-throughput DGGE analysis, and have successfully obtained the useful information of bacterial communities in different NJOF soils/sediments by Strategies 5.

Key words: Polymerase chain reaction (PCR), denaturing gradient gel electrophoresis (DGGE) analysis, bacterial diversity, soils/sediments, Northern Jiangsu Oil Field (NJOF).