4,501 SNPs consistent with transfer from Eagan (i e they were in

4,501 SNPs consistent with transfer from Eagan (i.e. they were in the same genome location as the Eagan SNPs identified above) were found in the Rd+EaganstrR transformants. We identified 202 SNPs that were common to all respective sequence reads, were not linked closely to other SNPs and were found in both Rd+EaganstrR and Rd+Eagan transformants obtained in control experiments using non-strR Eagan DNA as donor. We conclude that these SNPs were consistent with, and most likely explained by, errors within the reported Rd genome sequence published in 1995. Another possibility,

not mutually exclusive with sequencing errors, could be sequence drift in our laboratory strain (RM118) when compared to the sequenced isolate (Rd KW20). This level of error is similar to the several hundred SNPs reported upon re-sequencing of strain Rd by other investigators

GW786034 in vitro SHP099 chemical structure [17] and comparable with the 243 discrepancies found between the Ro-3306 original 1997 E. coli strain MG1655 genome sequence [19] and the 2006 re-sequencing [20] of the same strain. Figure 4 Frequency of Eaganstr R and Eagan SNPs in the Rd+Eaganstr R and Rd+Eagan transformants. Panel A; Location and frequency of EaganstrR specific SNPs plotted as estimated number of strains (y-axis) against location in RdKW20 genome sequence (x-axis) using SNPSeeker. MAQ was used to identify SNPs in the pooled sequences from 200 transformants. The location of the strR point mutation is indicated. Panel B; A magnified view of one region marked on Panel A showing a putative secondary transformation event. The extent of the chromosomal region involved with this predicted transformation event (13 kbp) is marked. Panel C; A magnified view of the primary transformation event from Panel A with the location of the strR point mutation marked. Panel D; The location and frequency of Eagan-specific SNPs in the genome of pooled Rd+Eagan transformants (200); Eagan unmarked (wild-type) genomic DNA was used as the donor. In the Rd+EaganstrR transformants, a large peak in SNP density centred on the site of the point mutation in rpoB conferring strR (Figure  4). Moving outwards from this central SNP peak,

the Eagan-specific SNPs decrease at a relatively constant rate such Flavopiridol (Alvocidib) that within 10 kbp of the strR mutation the frequency of strains containing Eagan-specific SNPs decreases at approximately 1 strain/100 bp. Across the 200 transformants, the region of the genome involved in recombination events centred on the strR locus would appear to span an arc of the genome over 80 kbp in size (Figure  4). Given that the strR locus can be at any location in the recombined block of DNA, this indicates a maximum size for the recombined block of at least 40 kbp. In addition to the intense peak centred on the strR conferring SNP, secondary small peaks of SNPs can be observed at other locations in the genome. These secondary peaks contain Eagan strain-specific SNPs at a frequency of approximately 0.

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