Patent ID: 6448002
Filing Date: 2002-09-10
Classification: C12N,C12Q,Y10S

Abstract:
A method for the detection of mutations in the DNA sequence of a ki-ras oncogene in DNA, comprising the steps ofa) incubating stool samples containing cells with a solution of activated charcoal to isolate the cells, b) lysing the cells with a buffer solution, wherein the buffer solution contains at least one chaotropic salt, to obtain a lysate; containing DNA, c) incubating the lysate in a washing buffer with a mineral carrier for binding the DNA to isolate the DNA from the lysate, d) separating the mineral carrier from the lysate by centrifugation, e) washing the DNA, which is bound to the mineral carrier, f) extracting the DNA by incubating the mineral carrier with a buffer of a low ionic strength, g) purifying the DNA with the buffer solution containing at least one chaotropic salt, h) incubating the buffer solution of g) containing the DNA with the mineral carrier to bind the DNA, i) separating the buffer solution of h from the mineral carrier by centrifuging, j) washing the mineral carrier to which the DNA is bound in the washing buffer, k) eluting the DNA from the mineral carrier with a second buffer of low ionic strength, l) amplifying the DNA with a primer pair and coupling the amplification reaction product to a solid phase, m) incubating the amplification reaction product with oligonucleotides that detect mutations of th ki-rass oncigene. n) hybridizing the amplification reaction product to the oligonucleotides, thereby detecting mutations in the DNA sequence of a ki-ras oncogene in the DNA.