Patent ID: 6607899
Filing Date: 2003-08-19
Classification: A61K,C07K,C12N,C12Q

Abstract:
A method of amplifying and isolating a nucleic acid molecule of interest from a mixture of nucleic acid molecules, comprising:(i) providing a recombinant nucleic acid library with a heterogeneous population of methylated, circular nucleic acid molecules as template molecules; (ii) annealing a first and a second primer to complementary strands of the circular nucleic acid molecule, to produce an annealed mixture, wherein the two primers extend in opposite directions relative to each other during polymerase chain reaction; the 5â€² ends of the two primers are adjacent to each other; and wherein each primer is identical in sequence to its corresponding sequence in the nucleic acid molecule of interest; (iii) subjecting the annealed mixture to polymerase chain reaction, thereby producing an amplified mixture containing linear amplicons; (iv) digesting the amplified mixture with an enzyme that selectively cleaves methylated nucleic acid, thereby eliminating the template molecules and enriching the nucleic acid molecule of interest; and (v) isolating the nucleic acid molecule of interest.