Quantitative polymerase chain reaction is also known as real-time fluorescence quantitative PCR. In this PCR technique, it gives the percentage of DNA in the sample. In the above RT PCR techniques, it is very cumbersome to detect the amount of DNA or RNA. But with qPCR technology, it can be quantified.
PCR Vs QPCR
Standard PCR can amplify up to 2000 nucleotides of DNA at a time. In contrast, QPCR can amplify and quantify DNA samples. Here, quantification is to measure how much DNA is in the sample.
PCR is based on the principle of DNA amplification through primer design and annealing. And QPCR works by using fluorescent probes and dyes. During DNA amplification or PCR amplification, the fluorescent probe emits fluorescence and can detect DNA samples.
Primer sets are used for standard polymerase chain reaction, while in QPCR, primers are labeled with fluorescent dyes for real-time polymerase chain reaction analysis. Primers used in standard polymerase chain reactions do not have any probe labels.
The standard polymerase chain reaction is accomplished in three steps: denaturation, annealing, and renaturation or extension. In the QPCR technique, through these steps, an additional exponential amplification stage is required to quantify the DNA.
For analysis by standard PCR, conventional gel electrophoresis uses PCR amplicons. In QPCR, however, the PCR machine records the results based on the fluorescence emitted by the dye.
In standard PCR, the bands of the amplicons are observed with the help of gel electrophoresis, based on the peaks of the bands. In QPCR, amplicons of different sizes can be observed in the peaks of QPCR.
PCR Vs RT PCR
In standard PCR, electrophoresis is carried out under the pressure of an electric current, so the amplicon band will migrate towards the positive pole. RT-PCR uses fluorescent dyes to detect DNA samples. The peak value of RT-PCR indicates the maximum amplification of the DNA sample.
In the above cases, all results of standard PCR are recorded after the process is completed. But with RT PCR, with fluorescence, results are quickly observed.
The amplification resolution of traditional polymerase chain reaction is lower, while the amplification resolution of RT PCR is higher.
It can be seen that the traditional polymerase chain reaction takes nearly 3 to 4 hours. Reverse transcription polymerase chain reaction (RT PCR) can amplify DNA in only 1 to 1.5 hours.