Method for identifying the source of an amplicon

Abstract

The present invention relates to a method for identifying the source of an amplicon, comprising: providing a plurality of pools of amplicons from different sources, wherein the amplicons from different sources are present in more than one pool, and wherein the amplicons in each pool are tagged with a unique pool-specific identifier; sequencing at least part of the amplicons that comprise the pool-specific identifiers; and assigning one or more of the amplicons to corresponding pools and/or sources using the pool-specific identifiers.

1 - 15 . (canceled) 
     
     
         16 . A physical map of Arabidopsis thaliana constructed de novo based on bacterial or yeast artificial chromosomes. 
     
     
         17 . A composition comprising a pooled library of bacterial or yeast artificial chromosomes, wherein the pooled library comprises a plurality of subpools obtained by three-dimensional pooling, and wherein each subpool comprises DNA fragments from a plurality of bacterial or yeast artificial chromosome clones. 
     
     
         18 . A method for the generation of a physical map of at least part of a genome comprising the steps of:
 (a) providing a sample DNA;   (b) generating a bacterial or yeast artificial chromosome clone bank wherein each artificial chromosome clone contains part of the sample DNA;   (c) combining the artificial chromosome clones in one or more pools, wherein each clone is present in more than one pool, to create a library;   (d) digesting the one or more pools with one or more restriction endonucleases to provide for a set of restriction fragments for each pool;   (e) 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;   (f) combining the adaptor-ligated restriction fragments;   (g) amplifying the adaptor-ligated restriction fragments of step (e) with at least one primer, which primer contains a pool-specific section corresponding to the pool-specific identifier section in the adaptor or contains a pool-specific identifier at the position of the degenerate identifier section, respectively, to provide amplified adaptor-ligated restriction fragments or amplicons;   (h) combining the amplicons in a set of combined amplicons;   (i) determining the sequence of at least the pool-specific identifier and part of the restriction fragment of the amplicons or set of combined amplicons;   (j) assigning the restriction fragment sequences determined in the amplicons of step (i) to the corresponding clones using the pool-specific identifiers;   (k) ordering the restriction fragments derived from the same clone to build a contig; and   (l) ordering the contigs of the clones of step (k) to thereby build a clone-contig and generate a physical map.