Blakesley et al. BMC Genomics 2010, 11:21 http://www.biomedcentral.com/1471-2164/11/21 Page 9 of 15 Figure 3 Gaps in comparative-grade finished BAC sequences from multiple genomic regions. The comparative-grade finished sequences of the 2,031 BACs summarized in Tables 2 and 3 were analyzed for the presence of uncaptured (orange bar) and captured (blue bar) gaps (see text for details). The numbers of captured and uncaptured gaps per Mb averaged across all BACs from each indicated genomic region (A) or species (B) are indicated. Each error bar represents the 95% confidence interval. In A, the data for three additional ENCODE pilot project regions (ENr231, ENr232, and ENr333) are shown for comparison because of the notably high frequency of gaps in their sequences; however, there were not sufficient numbers of sequenced BACs from these regions to qualify for inclusion in the second data set (see text for details). approach has not been particularly successful for some species (e.g., platypus). In these cases, the sequence assemblies frequently contain too many contigs that cannot be unequivocally ordered even with fosmid- derived read-pairs (in part because the fosmid insert is too large, with the resulting insert-end reads simply skipping over many contigs and gaps). As an alternative, we found that generating read pairs with a 10- kb insert plasmid library was more productive. Another approach that we have implemented involves the use of an alternate bacterial host strain for constructing the initial shotgun plasmid library. We rea- soned that uncaptured gaps might reflect regions of for- eign DNA that are ‘poisonous’ to the bacterial host, with subclones propagating such DNA in high copy- number plasmids either failing to grow or growing poorly [30-32]. Thus, a host that constrained plasmid copy number might reduce any bacterial growth inhibi- tion, yielding a more uniform distribution of sequence reads across the starting template. To test this idea, we selected three previously gener- ated BAC sequences (produced using our standard