A method for detecting a point mutation in a gene of interest using pcr

Abstract

The present invention relates to a novel method for the rapid and unambiguous detection of point mutations which, unlike existing methods, is not dependent on the conservation of nucleotides around the point mutation or mutations of interest. The present invention accomplishes this by providing a screening method in which a PCR is carried out on an extract of DNA from the gene of interest using a pair of PCR primers, the PCR product being digested by at least one restriction enzyme and the restriction digest analysed, the method characterised in that, (i) the 3' ends of the PCR primers are separated by 3 or fewer nucleotides, (ii) more than one mutation of interest is present in the 3 or fewer nucleotides between the 3' ends of the primers and (iii) the product of the PCR reaction is such that each mutation gives a different restriction profile when at least one restriction enzyme is used to digest the PCR product and this restriction profile is different from that of the wild type sequence.