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The microbial world

Mycobacterium tuberculosis. The microbial world. S. Cerevisiae (yeast). Metagenomics. Diversity Analysis 16sRNA sequencing Fingerprinting Hybridization. Last week: Extracted DNA Amplified 16s rRNA gene This week: Visualize PCR product (gel electrophoresis)

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The microbial world

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  1. Mycobacterium tuberculosis The microbial world S. Cerevisiae (yeast)

  2. Metagenomics Diversity Analysis 16sRNA sequencing Fingerprinting Hybridization • Last week: • Extracted DNA • Amplified 16s rRNA gene • This week: • Visualize PCR product (gel electrophoresis) • Functional analysis for antibiotic resistance http://wikieducator.org/images/4/4c/Metagenomics.jpg

  3. PCR mixture requirements DNA Primers MgCl2 Buffer dNTPs Taq DNA polymerase

  4. Metagenomics Gel Electrophoresis

  5. Metagenomics Functional Analysis Are there genes producing proteins that lead to antibiotic resistance?

  6. Metagenomics Bacteria lawn Disc containing antibiotic Clearing zone Similar to Kirby-Bauer disk diffusion susceptibility test - standardized protocol to measure resistance or sensitivity of pathogens to potential antibiotic compounds.

  7. Metagenomics • Resistant colonies –things to consider • Distance from disc • Number Factors affecting size of clearing zone: Effectiveness of antibiotic Concentration of antibiotic Solubility of antibiotic Depth of agar Density of bacteria

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