Supported PCR: an efficient procedure to amplify sequences flanking a known DNA segment

George N. Rudenko, Caius M.T. Rommens, H. John J. Nijkamp, Jacques Hille

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Abstract

We describe a novel modification of the polymerase chain reaction for efficient in vitro amplification of genomic DNA sequences flanking short stretches of known sequence. The technique utilizes a target enrichment step, based on the selective isolation of biotinylated fragments from the bulk of genomic DNA on streptavidin-containing support. Subsequently, following ligation with a second universal linker primer, the selected fragments can be amplified to amounts suitable for further molecular studies. The procedure has been applied to recover T-DNA flanking sequences in transgenic tomato plants which could subsequently be used to assign the positions of T-DNA to the molecular map of tomato. The method called supported PCR (sPCR) is a simple and efficient alternative to techniques used in the isolation of specific sequences flanking a known DNA segment.
Original languageEnglish
Number of pages6
JournalPlant Molecular Biology
Volume21
Issue number4
DOIs
Publication statusPublished - 1993

Keywords

  • transposition
  • T-DNA tagging
  • target enrichment
  • supported polymerase chain reaction (sPCR)

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