Detection of alterations in a gene by long range PCR using human mobile elements

摘要

A method for detecting the presence absence of an alteration in a gene of interest, comprises: a) subjecting a test sample comprising genomic DNA to long range polymerase chain reaction amplification of target DNA, wherein the long range polymerase chain reaction comprises use of a polymerase having 3' to 5' exonuclease activity, wherein the target DNA comprises a human mobile element of interest and wherein the long range polymerase chain reaction amplification utilises a primer which comprises all of the human mobile element of interest or is within or overlaps the human mobile element of interest, such that if the test sample comprises genomic DNA comprising the alteration in the gene of interest, PCR products are formed, and if the test sample does not comprise genomic DNA comprising the alteration in the gene of interest, no PCR products are formed; and b) detecting the presence or absence of the PCR products, wherein the presence of PCR products is indicative of the presence of the alteration in the gene of interest in the test sample, and the absence of PCR products is indicative of the absence of the alteration in the gene of interest in the test sample. Preferably the gene of interest is the C4A gene. Also described are methods wherein differences in size of the PCR products are assessed, methods of determining whether an individual is at risk of developing a disease associated with an alteration in a gene of interest and methods of determining C4A deletion genotype of an individual.