Sequence-specific detection of PCR-amplified DNA by restriction enzyme release of hybrids

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• 作者：Estrada ; Georgina ; Colin ; Laurence ; Gaytan ; Paul ; Alagó ; n ; Alejandro ; Lizardi ; Paul M.
• 刊名：Molecular and Cellular Probes
• 出版年：1996
• 出版时间：June, 1996
• 年：1996
• 卷：10
• 期：3
• 页码：179-185
• 全文大小：207 K

文摘

We have developed a method to detect PCR amplicons that contain a restriction enzyme site. The method is called Polymerase Chain Reaction-Hybridization and Enzymatic Release Assay (PCR-HERA). The PCR product is hybridized to a synthetic oligonucleotide that is immobilized on a microtitre plate. Specific cleavage of the resulting hybrid by a restriction endonuclease liberates a fluorescein labeled arm of the immobilized DNA. We demonstrate the use of PCR-HERA to detect species-specific sequences of extra-chromosomal, ribosomal DNA, ofEntamoeba histolytica. This highly specific assay does not require any washing steps and can be easily automated.