Sep 09, 2009 by Ph.D. Rafael Montalvo Rodríguez

Students will be exposed to fundamental molecular microbiology methods used for the characterization of bacterial isolates and communities, complement phenotype-based taxonomy with basic molecular methods and reinforce a research oriented approach to the characterization of unknown isolates and microbial communities. Students will recognize advantages and limitations of the experimental methods used and how these affect the interpretation of results. Most of the exercises currently offered in the course are limited to basic staining procedures, conventional light microscopy and culture-based techniques. Bacterial characterization is based solely on traditional biochemical tests with limited exposure to modern, molecular tools, inconsistent with the increasing prevalence of DNA-based methods in modern microbiology. Furthermore, recent recommendations from the FDA are to implement DNA-based identification of microbes of relevance in the sterile drug manufacturing setting and to increase the frequency of DNA amplification methods in clinical diagnosis of infectious diseases. In this module the students will do DNA extraction from Gram-positive and Gram-negative bacteria using a bead beating-based protocol with a standard chemical lysis method. The genomic material can be used for performing PCR genomic fingerprinting of a collection of isolates in order to discriminate among strains of the same species. Isolates will be provided from real research projects. The students will be introduced to the concepts of molecular characterization of bacteria and polymerase chain reaction (PCR). The small subunit (16S) rRNA genes will be amplified from DNA extracted in Session 2, hence conducting genomic fingerprinting (RFLP) allowing the characterization of bacterial cultures based on restriction analysis. A questionnaire on research content, skills, tools and techniques will be offered on the first day of lab and after completing the lab module.

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