Lecture 12 - Selected Molecular Biology Techniques Present...

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Selected Molecular Biology Techniques Present in Primary Scientific Literature
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Overview The assays/techniques presented herein take advantage of interactions between biological and non-biological molecules to afford researchers a deeper understanding of some aspect in Molecular Biology. These are the techniques employed everyday in molecular biology labs around the world and are infinitely useful in answering key questions concerning the mechanical mystery surrounding the science of life After the name of each technique there is found a certain number of “hits”, that is, the number of scientific journal articles found in a general search of PubMed using only the name of the technique
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Techniques in this Presentation 1. Reverse Transcriptase PCR 2. Real Time PCR 3. Western, Southern and Northern Blots 4. Gel Mobility Shift Analysis 5. MicroArray
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1. Reverse Transcriptase PCR (44610 hits) What it is: PCR performed with Reverse Transcriptase This allows RNA to be used as the “template” material and results in the production of DNA amplicons What it is often used for in lab: Turning isolated cellular mRNA into cDNA ( complimentary DNA ) to allow for later sequencing/identification, cloning, etc Special Considerations: As with any kind of PCR, it is much more useful if you know the sequence of the gene of interest (allows you to use unique primers) Usually not performed as an end unto itself; almost always coupled with some other assay (N or S blotting, Real Time PCR, etc)
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2. Real Time PCR ( 32217 hits) What it is: PCR performed in a dual thermocycler – spectrophotometer with primers that fluoresce once they are used by DNA Pol What it is often used for in lab: determining the actual amount of certain mRNAs present in a cell by performing Real Time PCR with Reverse Transcriptase on cellular mRNA isolate Special Considerations: much more specific AND sensitive than blotting techniques (N or S) even when the same primers are used, and can assay multiple genes simultaneously
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Lecture 12 - Selected Molecular Biology Techniques Present...

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