HLSC 3P97 Lecture Notes - Lecture 7: Reverse Transcription Polymerase Chain Reaction, Scanning Electron Microscope, Pneumocystis Jirovecii
2 Apr 2018
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Contrast: ability to distinguish between sample and background. Direct (wet) examination: cells are live under the microscope, no details. The light that is available to the specimen is diffused, background is dark, specimen is light. Fluorescent dye on the specimen will be absorbed. Specimen gets beam of electrons fired at it, examining the whole surface or sample --> 3d. Pcr tells us what genes are present and if certain genes are not present. Denature the dna to unzip the strands and provide access to the nucleotides. Add primers to the dna --> will bind to complementary ssequences. Lower temp and anneal primers 5" to 3". Taq polymerase attaches primers, and builds new strands. Rt-pcr suspect starting with specimen that has an rna genome. Taq polymerase is heat stable, so this step is done at a higher temp. Uses reverse transcriptase which translates rna to dna so that we can use. Real-time pcr quantitative tells us how heavy the infection is.
