Figure 3.

Rec8 Activated in Fetal Oocytes Builds Cohesive Structures

(A) Mating scheme to obtain female F1 offspring of the required genotype Rec8^TEV/TEV(Tg)Stop/Rec8-Myc (Tg)Dppa3-MCM-P.

(B) Activation of Rec8-Myc during meiotic S phase in fetal oocytes. Pregnant Rec8^TEV/TEV(Tg)Stop/Rec8-Myc females are injected with 4-OHT on embryonic day E10.5 to induce deletion by Dppa3-MCM-P in embryos. Oocytes are isolated from F1 females with the appropriate genotype. Green, meiotic DNA replication; beige, homologous recombination; pink, dictyate stage.

(C and D) Metaphase I chromosome spread showing localization of Rec8-Myc to bivalent chromosomes in oocytes from (C) wild-type and (D) F1 Rec8^TEV/TEV(Tg)Stop/Rec8-Myc (Tg)Dppa3-MCM-P female; oocytes analyzed n = 9 and n = 7, respectively. Centromeres are marked by anti-centromere antibody (ACA). Scale bar, 20 μm.

(E and F) Representative still images of oocytes isolated from F1 Rec8^TEV/TEV(Tg)Stop/Rec8-Myc (Tg)Dppa3-MCM-P females microinjected with mRNA encoding H2B-mCherry, CenpB-EGFP, and TEV protease.

(E) Bivalents are converted to chromatids in cells without Stop cassette deletion; n = 17 oocytes.

(F) Bivalents are retained in oocytes with successful deletion of the Stop cassette; n = 5 oocytes. Genotype of single cells was confirmed after live-cell imaging. Oocytes were obtained from >3 females. Scale bar, 20 μm.

See also Figure S3 and Table S1.