Figure 5.

AAV8-Lhcgr restores fertility and produces fertile offspring

(A) Representative images of oocytes, fertilization, one-cell embryos, and two-cell embryos. Scale bar: 50 μm.

(B) Offspring (F1) derived from AAV8-Lhcgr-treated Lhcgr^−/− male mice.

(C) Genotyping of the offspring derived from AAV8-Lhcgr-treated Lhcgr^−/− males and Lhcgr^+/+ females. The amplified wild-type (wt) DNA is 294 bp, while the mutant (mt) DNA fragment is 804 bp.

(D) PCR analysis of AAV8-Lhcgr integration in the genomes of offspring. CAG promoter- and Lhcgr-specific primers were used. As a control, tail DNA from Lhcgr^+/− mice was spiked with viral particles representing 0.1 and 1 copies of the viral genome.

(E) Southern blot analysis of DNA samples from offspring mice hybridized with AAV vector. Controls represent viral DNA in amounts equivalent to one and 10 copies of viral DNA per diploid genome. DNA was digested with the indicated restriction enzymes.

(F) Mating scheme used to produce the second-generation (F2) mice.

(G) Male F1 mice were used to produce F2 by mating with Lhcgr^+/+ females.

(H and I) Continuous breeding assay starting at 6 weeks of age, showing numbers of litters (H) and pups per litter (I) between F1 males and Lhcgr^+/− males within 4 months (n = 4).

(J) Mating scheme used to produce F2 mice.

(K) Female F1 mice were used to produce F2 pups by mating with Lhcgr^+/+ males.

(L and M) Continuous breeding assay starting at 6 weeks of age, showing numbers of litters (L) and pups per litter (M) between F1 females and Lhcgr^+/− females within 4 months (n = 4). Data are represented by boxplots, and whiskers show the minimum to maximum values. ns, not significant.