Patent ID: 8071310

Claim:
A method for the identification of an insertion associated with a gene or sequence of interest in a member of a transposon population, comprising the steps of: (a) isolating, individually or in pools, genomic DNA of the transposon population; (b) optionally, pooling the DNA obtained in step (a); (c) restricting the DNA using one or more restriction endonucleases and ligating adaptors to the restriction fragments, thereby preparing adaptor-ligated restriction fragments; (d) amplifying the adaptor-ligated restriction fragments with a pair of primers, whereby one of the primers comprises a section that is complementary or capable of hybridizing to part of a transposon sequence and further comprises a sequencing primer binding site, wherein the other of the primers is at least complementary to the adaptor, wherein one or both primers comprise a tag; (e) optionally, pooling the amplification products of step (d) to create a library of amplification products; (f) optionally, fragmenting the amplification products in the library to create amplification library product fragments; (g) determining the nucleotide sequence of the fragments of (d), (e) or (f) using high throughput sequencing; (h) optionally, trimming the sequence of the fragments in silico to remove any adaptor and/or transposon related sequence information; (i) identifying one or more fragments of step (g) or (h) that are capable of aligning with nucleotide sequences from a database, thereby correlating the identified fragments with a gene or phenotype of interest represented in the database; (j) identifying members of the transposon population containing the fragment or fragments of step (i); (k) optionally, designing a probe or PCR primer pair based on the fragments of step (i) and using said probe or PCR primer to confirm transposon insertion in the gene of interest in the genome of the member or members identified in (j).