common in the gp120 coding portion than in the gp41 region. The probability that breakpoints were more or less clustered across env than could be accounted for by chance (given the null hypothesis that breakpoint positions occur randomly) was determined by a permutation test ( Figure 2B , bottom panel). Six major recombination-prone or ''hot'' regions (shaded light blue areas in Figure 2B ) could be defined as env regions where breakpoint clusters were bounded by statistically significant breakpoint ''cold spot'' (p,0.05). Each of the six identified breakpoint clusters contained at least one breakpoint cluster that constituted a statistically significant recombination ''hot-spot'' (p,0.01). While these recombination-prone regions covered only slightly more than half of the whole gene (55.3%), they included 81.6% of all the breakpoints (337/413) mapped. These six hot regions are areas where recombination occurs preferentially during HIV replication, irrespective of the parental strains involved.