History
Since its introduction in 1977, Northern blot had been used extensively for RNA quantification despite its shortcomings of: (a) being time-consuming, (b) requiring a large quantity of RNA for detection, and (c) being quantitatively inaccurate in the low abundance of RNA content. However, the discovery of reverse transcriptase during the study of viral replication of genetic material led to the development of RT-PCR which has since displaced Northern blot as the method of choice for RNA detection and quantification.
RT-PCR has risen to become the benchmark technology for the detection and/or comparison of RNA levels for several reasons: (a) it does not require post PCR processing, (b) a wide range (>10^7 fold) of RNA abundance can be measured, and (c) it provides insight into both qualitative and quantitative data. Due to its simplicity, specificity and sensitivity, RT-PCR is used in a wide range of applications from experiments as simple as quantification of yeast cells in wine to more complex uses as diagnostic tools for detecting infectious agents such as the avian flu virus.
However, RT-PCR is not without a major flaw of its own. The exponential growth of the reverse transcribed complementary DNA (cDNA) during the multiple cycles of PCR produces inaccurate end point quantification due to the difficulty in maintaining linearity. In order to provide accurate detection and quantification of RNA content in a sample, qRT-PCR was developed using fluorescence-based modification to monitor the amplification products during each cycle of PCR.
Read more about this topic: Reverse Transcription Polymerase Chain Reaction
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“It is the true office of history to represent the events themselves, together with the counsels, and to leave the observations and conclusions thereupon to the liberty and faculty of every man’s judgement.”
—Francis Bacon (1561–1626)
“The thing that struck me forcefully was the feeling of great age about the place. Standing on that old parade ground, which is now a cricket field, I could feel the dead generations crowding me. Here was the oldest settlement of freedmen in the Western world, no doubt. Men who had thrown off the bands of slavery by their own courage and ingenuity. The courage and daring of the Maroons strike like a purple beam across the history of Jamaica.”
—Zora Neale Hurston (1891–1960)
“What is most interesting and valuable in it, however, is not the materials for the history of Pontiac, or Braddock, or the Northwest, which it furnishes; not the annals of the country, but the natural facts, or perennials, which are ever without date. When out of history the truth shall be extracted, it will have shed its dates like withered leaves.”
—Henry David Thoreau (1817–1862)