To statistically test for the presence of recombination hot and coldspots in the experimentally determined recombination breakpoint distributions we used a modification of a permutation test described previously [44] . Unlike in analyses of natural recombinants, the breakpoint positions approximated in our experimental procedure were not subject to biases introduced by underlying degrees of parental sequence nucleotide variability and patchiness of parental sequence sampling. Rather than explicitly accounting for these biases when placing randomised recombination breakpoints as in the permutation test described by Heath et al. [44] , our modification of the test involved the completely randomised placement of recombination breakpoints. The test essentially involved the randomised recreation of 10,000 versions of our real dataset with each version containing exactly the same number of breakpoints between the same 17 parental sequence pairs observed in the real dataset. From breakpoint distributions determined for each of these 10,000 randomised datasets we were able to work out confidence intervals for expected breakpoint density variation given the completely random occurrence of recombination. For simulating the distribution of 133 breakpoints based on the combined effects of (i) the mechanistic recombination rate and (ii) selection for functional recombinants, local recombination rate data used to generate the graph in Figure 2B were first multiplied by the respective functionality scores given in Figure 3 for each corresponding region, yielding ''functionality corrected'' rates for each region. Once the expected breakpoint distribution of 133 unique recombinants determined by this method, the number of breakpoints present in a 50 nucleotides rolling window, sliding with a 10 nucleotides increment was calculated and plotted (in Figure 5B ) as function of the position along the gene. Deviations from expected degrees of breakpoint clustering given the null hypothesis of random breakpoint locations, was tested using the same modification of the Heath et al., [44] permutation test detailed above.