Fig. 8. Quantitative analysis of OTD and OTX2 proteins, otd², otd^2FL and Otx2^2c mRNAs and their stability in ES cells. (A) Western blot analysis of extracts from HeLa cells, HeLa cells transfected with an otd-expressing vector, and 7.5 d.p.c. wild-type, otd²/+ and otd^2FL/+ embryos. (B) Western blot analysis on extracts from 7.5 d.p.c wild-type, Otx2^+/– and Otx2^2c/Otx2^2c embryos. (C,D) Coomassie Blue staining of the gels in A and B. (E) otd², otd^2FL and Otx2^2c cytoplasmic and nuclear mRNA levels were determined in otd²/+, otd^2FL/+ and Otx2^2c/+ embryos and are reported as percentages of the Otx2 mRNA. otd² mRNA was also analysed at 7.5 d.p.c. Note that while the otd² cytoplasmic mRNA is considerably reduced, the otd² nuclear mRNA has accumulated significantly. (F) Stability of otd², otd^2FL and Otx2^2c mRNA is comparable to that of Otx2 mRNA in actinomycin D experiments performed on heterozygous ES cell lines.

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