METHOD OF DETECTING METHYLATION

摘要

The present invention relates to a method of screening for the presence of methylated DNA in a biological sample by using multiple methylation-sensitive restriction endonucleases. More particularly, the present invention relates to a method of quantitatively screening for the level of one or more methylated genes of interest without the requirement that an undigested internal reference sample is used as a point of reference against which relative quantification is calculated. The present invention is useful in a range of applications including, but not limited to, providing a simpler and more accurate means to determine DNA methylation status, such as in the context of diagnosing or monitoring conditions characterised by changes to DNA methylation.

主权项

1. A method of quantitatively screening for the methylation of a DNA region of interest in a biological sample, said method comprising:
(i) contacting DNA from said biological sample with two or more methylation-sensitive restriction endonucleases and digesting said DNA at 4-6 pg of said DNA per unit of said endonucleases per hour; (ii) quantitatively amplifying the digested DNA sample of step (i) using one or more forward primers and one or more reverse primers, which primers are directed to DNA sequences flanking one or more methylation specific restriction endonuclease recognition sequence regions; and (iii) quantifying the level of said methylated DNA region of interest.