Grape (vitis vinifera L.) BAC library construction, preliminary STC analysis, and Identification of Clones Associated with Flavonoid and Stilbene biosynthesis
Tomkins, JP, Peterson, DG, Yang, TJ, Ablett, EF, Henry, RJ, Lee, LS, Waters, DLE & Wing, RA 2001, 'Grape (vitis vinifera L.) BAC library construction, preliminary STC analysis, and Identification of Clones Associated with Flavonoid and Stilbene biosynthesis', American Journal of Enology and Viticulture, vol. 52, no. , pp. 287-291.
We have constructed a grape BAC library using the cultivar Syrah. The library contains 55,296 clones stored in 144 384-well microtiter plates. A random sampling of 381 BACs indicated an average insert size of 144 kb with a range of 30 to 355 kb, and less than 4% of the clones do not contain inserts. Eighty-nine percent of BAC clones in the library have an average insert size greater than 100 kb. Based on a genome size of 483 Mb, library coverage is 16.5 haploid genome equivalents. Screening the BAC library colony filters with cpDNA sequences showed that contamination of the genomic library with chloroplast clones was low (1.5%). Library screening of an 11X coverage (2 BAC colony filters) with 12 cDNA probes corresponding to flavonoid and stilbene biosynthesis genes resulted in an average of 13 hits per probe (range = 1 to 27). To gain a glimpse into the grape genome and evaluate the library for sequence-tagged connector (STC) development, 768 BAC clones were end sequenced in both forward and reverse directions. The STCs were queried against the SWISS-PROT database and significant hits were sorted according to putative function.