Rice Genetics IV - IRRI books - International Rice Research Institute

1999). The development of multiple-enzyme BAC libraries allows for improved genomerepresentation. The rice BAC libraries consist of 36,864 and 55,296 clones forHindIII (vector: pBeloBAC11) and EcoRI (vector: pIndigoBAC536), respectively.The average insert size of the HindIII library was 129 kb and the EcoRI library 121kb. Approximately 5% of each library contains no inserts and organelle DNA (chloroplastand mitochondria) contamination. Based on a haploid genome size of 431 Mbfor rice (Arumuganathan and Earle 1991) and taking into account levels of emptyvectors and organelle DNA contamination, the coverage of the HindIII and EcoRIlibraries is approximately 10.6 and 15.0 haploid genome equivalents, respectively.Combined coverage for both libraries provides a total of at least 25 haploid genomeequivalents.Both BAC libraries (libraries, filters, and clones) are available at cost from theCUGI BAC/EST Resource Center (www.genome.clemson.edu).BAC fingerprint databaseTo develop a physical map of the rice genome, we attempted to generate unique restrictionfragment fingerprints from 36,864 BAC clones from each library (for methods,see Marra et al 1997). DNA was isolated from 74,496 BAC clones, digested withHindIII, run on high-resolution agarose gels, stained, and imaged. The HindIII fingerprintimages were then processed through IMAGE (Sulston et al 1989), whichtracks the lanes, calls the band, and creates band files. The band files were then inputinto FPC (Soderlund et al 2000), which assembles BAC clone fingerprints into contigsbased on the number of shared bands. Table 1 summarizes our preliminary results inTable 1. CUGI rice FPC project summary.Total Successful Clones Singleton No. of contigsin contig74,496 64,233 61,309 2,924 1,039(86.2%) (82.3%) (3.9%)% of clones706050403020100