DHPLC Technology as a High-throughput Detection of Mutations in a Durum Wheat TILLING Population
Poster Feb 02, 2015
Colasuonno P.1, Incerti O. 1, Lozito M.L. 1, Sbalzarini M. 2, Zaccagna P. 2, Papadimitriou S. 2, Blanco A. 1, Gadaleta A. 1
Durum wheat (Triticum turgidum ssp. durum) has a tetraploid genome (AABB genome) with genes in two homoeologous copies, a high proportion of repetitive DNA regions and a low gene density. Current tools for the discovery of sequence alterations in target genes rely on reverse genetic approaches such as the TILLING (Targeting Induced Local Lesions IN Genomes) technique. TILLING is based on the detection of chemical induced mutations in heteroduplex mainly using endonuclease (CelI enzyme), standard gel electrophoresis, fluorescent capillary electrophoresis and/or LI-COR techniques. Alternatively, DHPLC approach reveals the presence of genetic variation by the differential retention of the homo- and heteroduplex DNA molecules on ion-pair reversed-phase high-pressure liquid chromatography supported under partial denaturing conditions.
Despite the developments in conventional PCR, the complexity of multiplex Real Time PCR is still limited due to the lack of sufficient detection channels. To achieve high-end multiplexing capacity on standard Real Time PCR machines, Anapa Biotech has developed the MeltPlex® technology (see box on right).READ MORE