Complexity management of genomic DNA

摘要

The presently claimed invention provides for novel methods and kits for reducing the complexity of a nucleic acid sample by providing non-gel based methods for amplification of a subset of the sequences in a sample. In a preferred embodiment, amplification of a subset can be accomplished by digesting a sample with two or more restriction enzymes and ligating adaptors to the fragments so that only a subset of the fragments can be amplified. The invention further provides for analysis of the above amplified sample by hybridization to an array, which may be specifically designed to interrogate the desired fragments for particular characteristics, such as, for example, the to presence or absence of a polymorphism.

主权项

1. A method of amplifying fragments of a nucleic acid sample comprising:
fragmenting the nucleic acid sample using a first restriction enzyme and a second restriction enzyme, thereby producing a fragmented nucleic acid sample including double stranded nucleic acid fragments having a first end cut by the first restriction enzyme and a second end cut by the second restriction enzyme; ligating a first double-stranded adaptor to the first end of the double stranded nucleic acid fragments and a second double-stranded adaptor to the second end of the double stranded nucleic acid fragments, wherein one strand of the first adaptor comprises a 5′ overhang that provides a template for a primer binding site and one strand of the second adaptor comprises a 3′ overhang comprising a primer binding site, thereby producing ligated fragments comprising the 5′ overhang and the 3′ overhang, wherein the 5′ overhang and the 3′ overhang are present on the same strand of each of the ligated fragments comprising the 5′ overhang and the 3′ overhang; and exponentially amplifying the ligated fragments comprising the 5′ overhang and the 3′ overhang.