Method for genome sequencing using a sequence-based physical map
AssigneeKeygene N.V.
InventorsMichael Josephus Theresia Van Eijk, Taco Peter Jesse
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Abstract
The present invention relates to a high throughput method for the identification and detection of molecular markers wherein restriction fragments are generated and suitable adaptors comprising (sample-specific) identifiers are ligated. The adapter-ligated restriction fragments may be selectively amplified with adaptor compatible primers carrying selective nucleotides at their 3′ end. The amplified adapter-ligated restriction fragments are, at least partly, sequenced using high throughput sequencing methods and the sequence parts of the restriction fragments together with the sample-specific identifiers serve as molecular markers.
Claims22
A method for genome sequencing, comprising:
- (a) digesting artificial chromosome clones in a plurality of pools with one or more restriction endonucleases to provide for a set of restriction fragments for each pool, and wherein each clone is present in more than one pool;
- (b) ligating adaptors to one or both sides of the restriction fragments, wherein at least one adaptor contains a pool-specific identifier or a degenerate identifier section, respectively, to provide adaptor-ligated restriction fragments;
- (c) sequencing at least the pool-specific identifier and part of the restriction fragment;
- (d) assigning the restriction fragment sequences of step (c) to the corresponding clonal sources using the pool-specific identifiers;
- (e) generating a physical map by ordering the restriction fragment sequences;
- and
- (f) applying the physical map for assembling a genome sequence.
A method for genome sequencing, comprising:
- (a) ligating adaptors to one or both sides of restriction fragments obtained from digesting artificial chromosome clones in a plurality of pools, wherein each clone is present in more than one pool, with one or more restriction endonucleases, wherein at least one adaptor contains a pool-specific identifier or a degenerate identifier section, respectively, to provide adaptor-ligated restriction fragments;
- (b) sequencing at least the pool-specific identifier and part of the restriction fragment;
- (c) assigning the restriction fragment sequences of step (b) to the corresponding clonal sources using the pool-specific identifiers;
- (d) generating a physical map by ordering the restriction fragment sequences;
- and
- (f) applying the physical map for assembling a genome sequence.
Dependencies are inferred from the claim text.