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US 6,887,666 B1Granted
Methods for identification of identical nucleic acid fragments from different nucleic acid populations
AssigneeIntegragen
InventorJoerg Hager
Life of this patent
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Abstract
The present invention relates to genetic mapping of complex quantitative and qualitative traits. This invention more particularly relates to methods to identify identical DNA fragments from two different DNA sources. The method allows the amplification of the DNAs, their, labelling, and the separation of perfectly matched DNAs from imperfectly matched DNAs or from DNAs formed through hybridisation from the same source (e.g., homohybrids). The invention can be used to identify genes or gene mutations, which are relevant to pathological conditions or particular traits.
Claims14
A method comprising:
- (a) digesting separately nucleic acids from a mixture of at least two nucleic acid populations with at least one restriction enzyme;
- (b) ligating a blunt ended adaptor sequence to the restriction fragments resulting from the digestion in step (a);
- (c) amplifying adaptor-ligated restriction fragments generated in step (b) using an adaptor-specific primer to produce amplification products having different ends in respect to each population;
- (d) hybridizing the amplification products of step (c) from the different nucleic acid populations with each other to generate a mixture comprising homoduplexes and heteroduplexes;
- (e) eliminating blunt ended homoduplexes from heteroduplexes having forked ends by digesting the homoduplexes with an enzyme that specifically digests blunt ended double-stranded DNA fragments;
- (f) eliminating mismatched heteroduplexes by using mismatch repair enzymes;
- and
- (g) identifying, isolating or separating fully-matched heteroduplexes, thereby identifying, isolating or separating nucleic acid fragments that are identical between said at least two nucleic acid populations.
5 more elements
Dependencies are inferred from the claim text.