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:
“The mode of founding a college is, commonly, to get up a subscription of dollars and cents, and then, following blindly the principles of a division of labor to its extreme,—a principle which should never be followed but with circumspection,—to call in a contractor who makes this a subject of speculation,... and for these oversights successive generations have to pay.”
—Henry David Thoreau (1817–1862)
“In child rearing it would unquestionably be easier if a child were to do something because we say so. The authoritarian method does expedite things, but it does not produce independent functioning. If a child has not mastered the underlying principles of human interactions and merely conforms out of coercion or conditioning, he has no tools to use, no resources to apply in the next situation that confronts him.”
—Elaine Heffner (20th century)
“The proclamation and repetition of first principles is a constant feature of life in our democracy. Active adherence to these principles, however, has always been considered un-American. We recipients of the boon of liberty have always been ready, when faced with discomfort, to discard any and all first principles of liberty, and, further, to indict those who do not freely join with us in happily arrogating those principles.”
—David Mamet (b. 1947)