January 9-13, 2010
Town & Country Convention Center
San Diego, CA
Luca Dondini1 , Marco Adami1 , Orietta Lain2 , Federica Cattonaro4 , Remo Chiozzotto3 , Filippo Geuna3 , Stefano Tartarini1 , Daniele Bassi3 , Raffaele Testolin2 , Rosa Ye5 , David Mead5 , Cheng-Cang Wu5
Apricot (Prunus armeniaca L.) is one of the most important species of the stone fruit crops. A molecular genetic approach has been initiated to map putative regions associated with resistance and fruit quality in this species. BAC libraries have been employed for a wide variety of applications, such as physical mapping and map-based cloning, allowing the identification of genomic regions containing important loci for the discovery of new genes. Standard BAC library construction methods are based on partial restriction digestion of high molecular weight (HMW) genomic DNA. Because restriction enzyme sites are not evenly distributed across a genome, the resulting libraries are substantially biased, with numerous gaps which cannot be eliminated even by rising many fold the genome equivalent coverage. Gaps due to restriction enzyme bias are expected to be eliminated by novel methods to randomly shear HMW genomic DNA into very large fragments up to ~250 kb.
Here we describe the construction and preliminary characterization of an apricot BAC library using a random-shear approach coupled to the DNATerminator® End Repair reagents (Lucigen) to prepare inserts for efficient ligation. This library was constructed from the cultivar Lito, an important parent in apricot breeding programs and comprises 33,366 clones for an estimated 10X genome coverage. As a first application of the BAC library, we identified about 50 clones carrying SSR marker alleles from different regions of the genome tagged for Plum Pox virus (PPV) resistance and fruit quality, thus confirming the suitability of this library for gene identification and physical mapping of the apricot genome. The average clone size and degree of coverage will be also discussed.