Which of the following order of steps should be followed to produce gene-targeted animals?

• A. Inject female with PMSG; inject female with hCG; mate female with stud male; harvest embryo.
• B. Inject stud male with PMSG; inject female with hCG; mate female with dud male; harvest embryo.
• C. Inject stud male with PMSG; mate female with stud male; inject female with hCG; harvest embryo.
• D. Mate female with dud male; inject female with PMSG; inject female with hCG; harvest the embryo.

Answer: (A)

Maximizing embryo yield through controlled superovulation and timely fertilization is what this sequence targets. First, injecting the female with PMSG stimulates multiple follicle development, producing more oocytes available for fertilization. Then delivering hCG triggers ovulation so those mature oocytes are released in a synchronized window. Introducing a stud male at this point allows fertilization to occur, yielding embryos that can be harvested soon after. The embryo harvest should come after mating, once fertilization has occurred and embryos are at the appropriate early stage for downstream gene-targeting steps. Other sequences misplace actions: there is no role for PMSG in males, so injecting a stud male is inappropriate; hCG after or without ovulation would not synchronize ovulation; mating before ovulation would reduce fertilization success and embryo yield; and harvesting embryos before mating cannot yield fertilized embryos. Therefore this sequence best fits the standard protocol for producing gene-targeted animals.