A maize bacterial artificial chromosome (BAC) library from the European flint inbred lined F2

We report here the construction and characterisation of a BAC library from the maize flint inbred line F2, widely used in European maize breeding prog rams. The library contains 86,858 clones with an average insert size of app roximately 90 kb, giving approximately 3.2-times genome coverage. High-effi ciency BAC cloning was achieved through the use of a single size selection for the high-molecular-weight genomic DNA, and co-transformation of the lig ation with yeast tRNA to optimise transformation efficiency. Characterisati on of the library showed that less than 0.5% of the clones contained no ins erts, while 5.52% of clones consisted of chloroplast DNA. The library was g ridded onto 29 nylon filters in a double-spotted 8 x 8 array, and screened by hybridisation with a number of single-copy and gene-family probes. A 3-d imensional DNA pooling scheme was used to allow rapid PCR screening of the library based on primer pairs from simple sequence repeat (SSR) and express ed sequence tag (EST) markers. Positive clones were obtained in all hybridi sation and PCR screens carried out so far. Six BAC clones, which hybridised to a portion of the cloned Rp1-D rust resistance gene, were further charac terised and found to form contigs covering most of this complex resistance locus.