Yes, if you are going to register the litter you will have to DNA both boys, the mom, and all the pups. It can be pretty costly from what I have heard. Some breeders do this to get pups from 2 males for their own reasons. You then may have to pay for the DNA results to be read at $40 each reading. Then there is an additional fee of $200 in addition to regular registry. it seems pretty complicated. Here is the info for you:
American Kennel Club - Multiple-Sired Litter Registration