Figure 3. Restriction decreases integration length during inter-strain DNA transformation.
A) Distribution of SNPs and unique restriction sites used for mapping integration endpoints. The selective marker cat was integrated within gene cagH (triangle, point 0) and the 15 kbp surrounding DNA sequence contains approximately 600 SNPs (vertical bars on axis) that differentiate strains NSH57 and J99. Five restriction enzymes that have unique sites in either recipient (grey) or donor (black) DNA were used for mapping integration endpoints. The approximate positions of the coding regions are shown with black arrows. B), C), D). The length of integrated DNA segments was determined for 25 wild type clones (B) and 25 ΔΔhpyR(188III/CH4V) clones (C) transformed with homeologous gDNA, and for 18 wild type clones transformed with mosaic gDNA (D). For integration endpoints, the midpoint between informative SNPs was calculated and each integration event is represented with a horizontal bar ranked from larger to smaller. Integration events obtained from co-culture experiments are marked with (*). The size of the selective marker cat was not included in the overall integration length calculation. In D), wild type NSH57 was transformed with two distinct mosaic gDNA. Five of 18 transformants contained the entire 9–10kb donor DNA sequence (black) and 13 showed decreased integration length (grey). A t-test was used to compare the mean integration length of the ΔΔhpyR(188III/CH4V) mutant bacteria with that of wild type bacteria transformed with homeologous or mosaic gDNA.