Optimization of randomly amplified polymorphic DNA-polymerase chain reaction for molecular typing of Salmonella enterica serovar Typhi.
Optimization of the RAPD reaction for characterizing Salmonella enterica serovar Typhi strains was studied in order to ensure the reproducibility and the discriminatory power of this technique. Eight Salmonella serovar Typhi strains isolated from various regions in Brazil were examined for the fragment patterns produced using different concentrations of DNA template, primer, MgCl2 and Taq DNA polymerase. Using two different low stringency thermal cycle profiles, the RAPD fingerprints obtained were compared. A set of sixteen primers was evaluated for their ability to produce a high number of distinct fragments. We found that variations associated to all of the tested parameters modified the fingerprinting patterns. For the strains of Salmonella enterica serovar Typhi used in this experiment, we have defined a set of conditions for RAPD-PCR reaction, which result in a simple, fast and reproducible typing method.
Quintaes
Bianca Ramalho
BR
Laboratório de Zoonoses Bacterianas, Departamento de Bacteriologia, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, RJ, Brazil.
Leal
Nilma Cintra
NC
Reis
Eliane Moura Falavina
EM
Hofer
Ernesto
E
D015373:Bacterial Typing Techniques / Q000379:methods / Q000592:standards*; D001938:Brazil; D016172:DNA Fingerprinting / Q000379:methods; D004269:DNA, Bacterial / Q000032:analysis*; D006801:Humans; D019105:Random Amplified Polymorphic DNA Technique / Q000592:standards*; D015203:Reproducibility of Results; D012485:Salmonella typhi / Q000145:classification* / Q000235:genetics
D004269:DNA, Bacterial