¹ Clemson University Genomics Institute, Clemson, SC, 29634, USA, http://www.genome.clemson.edu

Two rice whole genome libraries were constructed with an average insert size of about 10kb (10kb library) to support the gap filling and finishing phases of the rice genome sequencing project. A series of plasmid vectors were constructed and one pCUGIblu21 was selected for library construction using HaeIII and Sau3AI partially digested genomic DNA. The HaeIII library contains 166,752 clones covering ~4.6x rice genome equivalents with an average insert size of 10.5kb. The Sau3AI library contains 138,960 clones covering 4.2x genome equivalents with an average insert size of 11.6kb. Both libraries were gridded onto 11 high-density filters in a 5x5 pattern, in duplicate. This 10kb library represents unbiased coverage throughout whole rice genome based on 12 OVERGO hybridizations (average 12 hits per each probe) and hybridization with telomeric repeat sequence (around 200 hits). Approximately 3% of the libraries are contaminated with chloroplast sequence. We are developing an efficient method to use this resource to fill relatively small gaps (up to 30kb) between sequenced BACs on rice chromosome 10. It will be an cost and time effective way to close the small sequencing gaps using one or two 10kb clones rather than using a 120kb BAC clone. We filled a 12,547bp gap using two 10kb clones and other gaps are in the process of being filled. A general procedure for gap filling and results will be presented in detail.