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Applications of Polymerase Chain Reaction (PCR) to Plant Genome Analysis

  • Majid R. Foolad
  • Siva Arulsekar
  • Raymond L. Rodriguez
Part of the Springer Lab Manual book series (SLM)

Abstract

The process of Polymerase Chain Reaction (PCR) was first described by Mullis et al. (1986), and it was for this invention that Dr. Kary Mullis received the 1993 Nobel Prize in Medicine and Physiology. PCR is a relatively simple process by which virtually unlimited copies of selected DNA fragments can be generated in a short period of time. This in vitro enzymatic amplification of specific DNA sequences involves three steps which are repeated a number of times (cycles): (1) DNA denaturation: the double-stranded template DNA (usually genomic DNA) is dissociated into single strands by heating the sample at 92–94 °C briefly. (2) Primer annealing: by lowering the temperature to 40–60 °C, two oligonucleotide primers (typically 18–22 bases in length) can anneal to regions on the single DNA strands that flank the target DNA sequence. The 3’ ends of each primer must face each other for the target DNA sequence to be amplified. (3) Primer extension: the 3’ ends of the oligonucleotide primers are extended toward each other with newly synthesized DNA. This new DNA is complementary to target DNA sequences. To reduce nonspecific annealing of primers to DNA, this step is usually performed at an elevated temperature (e.g., 72 °C) using a thermostable DNA polymerase (typically Taq DNA polymerase from Thermus aquaticus).

Keywords

Polymerase Chain Reaction Standard Polymerase Chain Reaction Downy Mildew Resistance Inverse Polymerase Chain Reaction Polymerase Chain Reaction Technology 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Springer-Verlag Berlin Heidelberg 1995

Authors and Affiliations

  • Majid R. Foolad
  • Siva Arulsekar
  • Raymond L. Rodriguez

There are no affiliations available

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