Principles
In RT-PCR, the RNA template is first converted into a complementary DNA using a reverse transcriptase. The cDNA is then used as a template for exponential amplification using PCR. RT-PCR is currently the most sensitive method of RNA detection available. The use of RT-PCR for the detection of RNA transcript has revolutionalized the study of gene expression in the following important ways:
- Made it theoretically possible to detect the transcripts of practically any gene
- Enabled sample amplification and eliminated the need for abundant starting material that one faces when using northern blot analysis
- Provided tolerance for RNA degradation as long as the RNA spanning the primer is intact
Despite its major advantages, RT-PCR is not without drawbacks. The extreme sensitivity of the technique can be a double edged sword since even the slightest DNA contamination can lead to undesirable results. Additionally, planning and design of quantification studies can be technically challenging due to the existence of numerous sources of variation including template concentration and amplification efficiency.
Read more about this topic: Reverse Transcription Polymerase Chain Reaction
Famous quotes containing the word principles:
“I cannot consent that my mortal body shall be laid in a repository prepared for an Emperor or a King—my republican feelings and principles forbid it—the simplicity of our system of government forbids it.”
—Andrew Jackson (1767–1845)
“Prohibition will work great injury to the cause of temperance. It is a species of intemperance within itself, for it goes beyond the bounds of reason in that it attempts to control a man’s appetite by legislation, and makes a crime out of things that are not crimes. A Prohibition law strikes a blow at the very principles upon which our government was founded.”
—Abraham Lincoln (1809–1865)
“I suppose that one of the psychological principles of advertising is to so hammer the name of your product into the mind of the timid buyer that when he is confronted with a brusk demand for an order he can’t think of anything else to say, whether he wants it or not.”
—Robert Benchley (1889–1945)