×

Method for high-throughput AFLP-based polymorphism detection

  • US 8,815,512 B2
  • Filed: 11/01/2012
  • Issued: 08/26/2014
  • Est. Priority Date: 12/22/2005
  • Status: Active Grant
First Claim
Patent Images

1. A method for the high throughput discovery, detection and genotyping of one or more genetic markers in one or more samples, comprising the steps of:

  • (a) providing DNA from one or more samples;

    (b) restricting the DNA with at least one restriction endonuclease to produce at least one restriction fragment comprising sticky remains of a restriction site;

    (c) ligating at least one adaptor to the at least one restriction fragment to produce at least one adaptor-ligated restriction fragment, wherein one end of the adaptor comprises an overhang that is compatible with the sticky remains of the restriction site of the restriction fragment;

    (d) amplifying by solution PCR the at least one adaptor-ligated restriction fragment with at least one primer pair to produce a library of at least one amplified adaptor-ligated double stranded restriction fragment, wherein the amplified adaptor-ligated restriction fragment comprises an identifier tag;

    (e) sequencing a pool of two or more libraries of tagged amplified adaptor-ligated restriction fragments using high throughput sequencing technology, wherein each library comprises a unique identifier tag, wherein the sequencing comprises amplifying by PCR on a solid support;

    (f) clustering the sequences per library, using the identifier tag;

    (g) identifying genetic markers within the library and/or between libraries;

    (h) determining dominant or co-dominant genotypes of the genetic markers in the two or more libraries.

View all claims
  • 0 Assignments
Timeline View
Assignment View
    ×
    ×