Effects of GC Clamps on RpoB-PCR-DGGE Fingerprint of Three Types of Food Borne Pathogens

doi:10.13560/j.cnki.biotech.bull.1985.2016.05.028

Abstract

Abstract: This work is to investigate the effect of different GC clamps and different positions of primers connected with GC clamps on the DGGE fingerprint of food borne pathogens. We synthesized 6 pairs of primers encoding RNA polymerase β subunit（rpoB 1-6）containing 3 different GC clamps（GC-1，GC-2 and GC-3）that linked to 5'endings of both forward and reverse primers，respectively. Then，rpoB-PCR-DGGE was used to analyze Vibrio parahaemolyticus（5 strains），Listeria monocytogenes（5 strains），and Salmonella spp.（3 strains）. The results of fingerprint analysis were compared with that of V3-PCR-DGGE and ERIC-PCR. GC-3 clamp linked with reverse primer presented the best discriminating effect，and the discriminating index of rpoB-PCR-DGGE reached 0.9 that was equal to ERIC-PCR and greater than V3-PCR-DGGE. In conclusion，both of the different GC clamp sequences and linked positions of primers with GC clamp affected the result of rpoB-PCR-DGGE fingerprint. Choosing or designing GC clamp without continuous guanine（G base）and linked with 5'ending of reverse primer will improve the detecting and typing effect of rpoB-PCR-DGGE for food borne pathogens.

Key words: Vibrio parahemolyticus, Listeria monocytogenes, Salmonella spp., GC clamps, rpoB-PCR-DGGE

LIAO Chao,ZHANG Zhao-huan,YAO Xin,XIE Jing,ZHANG Wei-jia,PAN Ying-jie,ZHAO Yong, ,. Effects of GC Clamps on RpoB-PCR-DGGE Fingerprint of Three Types of Food Borne Pathogens[J]. Biotechnology Bulletin, 2016, 32(5): 212-219.

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