PCR techniques

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Several techniques have have been derived from the basic polymerase chain reaction (PCR). Below is an overiew of important PCR methods linking to individual pages for detailed information.

  • nested PCR is 2 successive PCRs with the 2nd set of primers nested inside the 1st pair. It is used to reduce unspecific products.
  • multiplex PCR is a PCR with >1 primer pair run in a single reaction. It reduced material consumption but is hard to optimise. It is often used in mouse genotyping for example.
  • colony PCR is a PCR technique to detect DNA in bacterial colonies. Colonies are picked, lysed, and amplified by PCR. It is used to detect successful ligations or recombinations among large numbers of bacterial colonies.
  • RT-PCR (Reverse Transcription PCR) is essentially normal PCR preceded by a reverse transcription converting RNA to cDNA. It is used to indirectly detect/measure RNAs, e.g. messenger RNAs.
  • Q-PCR (quantitative PCR) is used to determine the quantity of DNA. There are 3 main methods: agarose gels, dsDNA dyes, and probes (more accurate last). Both dsDNA dyes and probes allow measurement while the PCR is running, i.e. in real-time, and are therefore often referred to as real-time PCR (see below).
    • real-time PCR is used to determine the quantity of DNA real-time and thus a subset of the qPCR methods. It uses fuorescent dyes, such as Sybr Green, or fluorophore-DNA probes, such as TaqMan, to measure the amount of amplification in real time. This is used to defer starting amount. Real-time PCR It is often confusingly abbreviated as RT PCR, which overlaps with reverse transcription PCR; QRT-PCR or RTQ-PCR are better acronyms.
      • real-time RT-PCR is a special type of real-time PCR with reverse transcribed RNA as template. Basically, RT + qPCR.


PCR on wikipedia

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