Topic 10, Reverse transcription and RT-PCR, microarrays.ppt.edu

Topic 10, Reverse transcription and RT-PCR, microarrays.ppt.edu

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Topic 10—Reverse transcription and RT-PCR, Microarrays
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Learning Objectives Be able to define the following terms: gene expression, cDNA, RT-PCR, Q-RT-PCR, reverse transcription, reverse transcriptase, microarray Be able to explain how cDNAs are produced and how they can be used as tools in molecular biology Be able to diagram how microarrays work and be able to interpret data Be able to differentiate between the methods used for PCR, RT-PCR, and Q-RT-PCR Be able to compare and contrast the various methods to detect and quantify gene expression After completing homework #2, be able to choose the most
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What is a gene? This used to be a straightforward question, the definition has now become more murky…. Classical definition: A DNA sequence that encodes a protein Updated definition: A DNA sequence that encodes a function at the level of RNA or protein (we now know of lots of RNAs that are never intended to be translated to protein)
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What is gene expression? Old definition: production of a protein: using DNA to make mRNA, then using mRNA to make protein New definition: Old definition still applies in most cases Some RNAs are functional without need for protein A cell can start the process of making a protein, then decide to change it’s mind and stop the
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Reverse Transcription Uses the enzyme reverse transcriptase (isolated from retroviruses) to convert RNA to cDNA cDNA = complementary DNA. A DNA molecule that is complementary and anti- parallel relative to the RNA used to produce it Reverse Transcriptase is an example of an “RNA-dependent DNA polymerase” mRNA 5’ UAGCUAGCUA 3’ cDNA 3’ ATCGATCGAT 5’
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Topic 10, Reverse transcription and RT-PCR, microarrays.ppt.edu

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