Construction of tomato genomic DNA libraries in a binary-BAC (BIBAC) vector

This is the first report of large insert genomic DNA libraries constructed in a binary-BAC (BIBAC) vector. Genomic DNA libraries containing approximat ely 4.6 haploid nuclear genomic equivalents were constructed for Lycopersic on esculentum (cv. Mogeor) and Lycopersicon pennellii (LA716) in the BIBAC2 vector. The L. esculentum library has an average insert size of 125 kb and is comprised of 42 272 individual colonies stored as frozen cultures in a 384-well format (108 plates). The L. pennellii library has an average inser t size of 90 kb and is comprised of 53 760 individual clones (140 384-well plates). In each of the libraries, it is estimated that 90% of the colonies contain genomic DNA inserts. The composition of the L. esculentum and L. p ennellii libraries was determined by analyzing a series of randomly selecte d clones. The L. esculentum library was surveyed for clones containing chlo roplast DNA (1.4%), mitochondrial DNA (0.012%) and repetitive DNA motifs. B IBAC clones that may contain a gene of interest can be identified from thes e libraries by colony hybridization with homologous or heterologous probes or by PCR pooling techniques. Once identified, BIBAC genomic DNA library cl ones are immediately suitable for Agrobacterium tumefaciens-mediated plant transformation.