Reverse Transcription Protocol - How Dna Sequences Are Copied

Reverse Transcription Protocol – How DNA Sequences Are Copied

By: - Science - August 25, 2011
reverse transcription protocol %E2%80%93 how dna sequences are copied

The reverse transcription PCR protocol is a series of steps for copying DNA in the molecularly biology laboratory. It uses polymerase chain reactions (PCR), a technique used by scientists in order to amplify a sequence of DNA. Amplification of DNA means creating many copies of that particular section of the DNA strand. Reverse transcription PCR protocol is slightly different from regular PCR in that the first step is for an RNA strand to be transcribed into its complementary DNA strand. This requires the enzyme reverse transcriptase and is done in a tube at a temperature between 40 degrees C and 50 degrees C. The PCR is then applied to the complementary DNA strand. Double stranded DNA is now denatured at higher temperatures (around 95 degrees C), causing the two strands to unwind from one another and begin new reactions as the temperature is lowered slowly. Two strands of nucleic acids called primers are lengthened using DNA polymerase and a copy is made. This occurs around 72 degrees C. In this way the strands are duplicated and multiply exponentially. The products should be analyzed via electrophoresis to ensure everything was done correctly in the reverse transcription protocol.

Because the reverse transcription protocol has such a high sensitivity, it can be used to detect things other tests cannot. It is still considered the most sensitive technique available for the detection of mRNA. Reverse transcription PCR protocol is so sensitive, in fact, that it can be used to measure RNA from a single cell.

The reverse transcription protocol is used in cloning. It is used for genetic disease diagnosis and for inserting the genes of a eukaryotic organism into a prokaryote. It is also useful for studying RNA viruses such as HIV. Advances in how HIV copies itself have lead to the development of antiviral drugs that have the potential to combat HIV. When HIV infects a healthy cell, the reverse transcriptase makes a copy of the single strand RNA and changes it into a double stranded DNA, which is now viral. The viral DNA will join up with the chromosomal DNA and allows transcription and translation to reproduce the virus, sending it all through the body.

By using reverse transcription protocol, scientists can help develop drugs known as reverse transcriptase inhibitors that will stop the HIV from copying itself. Ideally they can be used to treat not only HIV but cancer as well. This advanced DNA science may eventually help find cures to a variety of serious diseases and change the landscape of biomedicine forever.

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