Supported PCR: an efficient procedure to amplify sequences flanking a known DNA segment |
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Authors: | George N. Rudenko Caius M. T. Rommens H. John J. Nijkamp Jacques Hille |
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Affiliation: | (1) Department of Genetics, Free University, De Boelelaan 1087, 1081 HV Amsterdam, Netherlands |
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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. |
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Keywords: | supported polymerase chain reaction (sPCR) target enrichment T-DNA tagging transposition |
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