None of these assumptions made for the final TMR4A analysis.
Some of these assumptions are made for the “effective population size” estimates, which is why we discarded these studies after demonstrating the assumptions dramatically affect results. Relying upon these estimates was an error.
The main assumption made for TMR4A is the mutation rate, which we can directly verify both now and in the distant past. I suppose there also is the assumption that all humans descend from the same population.