Plant & Animal Genomes XIV Conference

January 14-18, 2006
Town & Country Convention Center
San Diego, CA

Using Restriction Enzymes As A Method Of Genome Filtering In Wheat

John P. Fellers

  USDA-ARS-PSERU 4008 Throckmorton Hall Manhattan, KS 66506

Bread wheat (Triticum aestivum) contains one of the largest genomes of the plant kingdom and is a significant challenge when considered for genome sequencing. Wheat is known to have gene-rich regions within the genome interspersed by vast amounts of repetitive DNA. It is understood that a filtering technique would be of significant benefit when targeting gene containing regions for sequencing. Previous work has indicated that the restriction enzyme, PstI can increase the presence of low copy DNA when used to digest genomic DNA. To evaluate this on a larger scale, genomic DNA of the bread wheat ‘Chinese Spring’ was digested with the methyl sensitive restriction enzymes, Sac1, Sal1, Pst1, and AatII. DNA fragments of >400 bp were subcloned into cloning vector, isolated, and single pass sequenced. Preliminary evidence indicates that this technique can enrich by more than 35%, depending on the enzyme used. The sequencing results will be evaluated for the presence of open reading frames, repetitive DNA and low copy DNA. The results will be reported here.