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The exponential amplification via reverse transcription polymerase chain reaction provides for a highly sensitive technique in which a very low copy number of RNA molecules can be detected. RT-PCR is widely used in the diagnosis of genetic diseases and, semiquantitatively, in the determination of the abundance of specific different RNA ...
Reverse transcription loop-mediated isothermal amplification (RT-LAMP) is a one step nucleic acid amplification method to multiply specific sequences of RNA. It is used to diagnose infectious disease caused by RNA viruses. [1] It combines LAMP [2] DNA-detection with reverse transcription, making cDNA from RNA before running the reaction. [3]
A reverse transcriptase (RT) is an enzyme used to convert RNA genome to DNA, a process termed reverse transcription.Reverse transcriptases are used by viruses such as HIV, COVID-19, and hepatitis B to replicate their genomes, by retrotransposon mobile genetic elements to proliferate within the host genome, and by eukaryotic cells to extend the telomeres at the ends of their linear chromosomes.
By adding a reverse transcriptase enzyme to an RPA reaction, it can detect RNA as well as DNA, without the need for a separate step to produce cDNA. [2] [3] [4] Because it is isothermal, RPA can use much simpler equipment than PCR, which requires a thermal cycler. Operating best at temperatures of 37–42 °C and still working, albeit more ...
Although RNA can also be amplified by PCR using a reverse transcriptase (in order to synthesize a complementary DNA strand as a template), NASBA's main advantage is that it works under isothermal conditions – usually at a constant temperature of 41 °C or two different temperatures, depending on the primers and enzymes used. Even when two ...
The mRNA of an input sample (e.g. a tumour) is isolated and a reverse transcriptase and biotinylated primers are used to synthesize cDNA from mRNA. The cDNA is bound to Streptavidin beads via interaction with the biotin attached to the primers, and is then cleaved using a restriction endonuclease called an anchoring enzyme (AE). The location of ...
Template-switching polymerase chain reaction (TS-PCR) is a method of reverse transcription and polymerase chain reaction (PCR) amplification that relies on a natural PCR primer sequence at the polyadenylation site, also known as the poly(A) tail, and adds a second primer through the activity of murine leukemia virus reverse transcriptase. [1]
The protocols for 5' or 3' RACES differ slightly. 5' RACE-PCR begins using mRNA as a template for a first round of cDNA synthesis (or reverse transcription) reaction using an anti-sense (reverse) oligonucleotide primer that recognizes a known sequence in the middle of the gene of interest; the primer is called a gene specific primer (GSP). The ...
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