So in theory you can breed DUs twice, but in reality there's generally no point doing that - it really only has disadvantages. I don't know anyone who actually breeds them twice, honestly. The only exception would be if for some reason (lore etc I guess?) you specifically wanted your litter to have two separate fathers. Otherwise I'd strongly recommend only breeding your DUs once.

The reason for that is that the second breeding (whether it's the same stud again, or a different stud) effectively overrides half the cubs generated from the first stud. So if on the first breeding you use items like a lion scrotum to try to pass down her mutation, or gnawrocks to pass markings, or anything else, then half those cubs are deleted on the second breeding and generated fresh. So unless you use double the items to effectively do the same thing all over again on the second breeding, half the cubs born aren't actually benefitting from the items used at all.

And even if you do use another set of items - and don't mind the waste of that - you could still potentially be deleting mutated/valuable cubs from the first breeding when they get overridden on the second breeding; you'd never know if the cubs you would have got from the first breeding would have been better...

The first breeding determines the number of cubs, so there's no benefit to breeding her again; a second breeding doesn't increase the amount of cubs or anything like that.

By far the best strategy imo is just to breed DUs once, like normal, and throw all your items at that one breeding. Especially a buffalo scrotum if possible - since that's the way to maximise your cubs, at either 6 or 8 for a DU litter. Or a grain of paradise at minimum, since that would still guarantee at least 4. That way no items are wasted and you aren't deleting potentially valuable cubs :)

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