expert corner
DIGITAL PCR-AN EMERGING DISRUPTIVE TECHNOLOGY FOR NUCLEIC ACID REPLICATION
With a higher degree of accuracy and reproducibility in the amplification and analysis of nucleic acids like DNA, RNA and cDNA, this technology is highly promising as compared to the existing forms of PCR
Developed in 1983, the Nobel Prize winning Polymerase Chain Reaction( PCR) has been one of the most commonly used technologies across life science laboratories. Primarily used for amplifying DNA, its application spans cloning, sequencing, analysis of genes, diagnosis and more. Traditional PCR is an end-point analysis. The PCR amplified product is detected by agarose gel electrophoresis making only a semi-quantitative analysis possible. A major drawback with PCR has been that
the quantification of the starting material was never possible. With rapid developments in molecular biology, the need for quantification of DNA in the PCR reaction has been growing.
The technique further improved such that quantitative estimation of DNA became possible and it became today’ s gold standard for measuring specific amounts of DNA in a sample. Real-time or Quantitative PCR( qPCR), is a very powerful and sensitive technique with a broader range of applications compared to PCR, including SNP detection, viral load detection, siRNA / miRNA / RNAi analysis etc.
qPCR utilizes specific complementary oligonucleotide primers that amplify the target of interest along with a fluorescent dye which tracks the progress of the reaction. The reaction requires comparison with a standard curve so as to arrive at the exact concentration of the amplified product. Boehringer Mannheim developed and commercialized first real time PCR. However, qPCR has some limitations such as the need for reference standards and has
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BioVoiceNews | July 2016