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. 2003 Jan 27;100(4):1844–1848. doi: 10.1073/pnas.0437912100

Figure 3.

Figure 3

Generation of GFP knockdown mice. (a) Decreased fluorescence of GFP in blastocytes transduced with LV-siGFP (♂, filled green depicts GFP transgenic male mouse; ♀, wild-type female). (b) F0 animals were analyzed by PCR of genomic DNA using U3 primers. The U3 primers amplified the H1-siGFP cassette cloned in the U3 region in the LTR. The siGFP-positive F0 mice (F0-2 + F0-4) are indicated in red. (c) PCR of genomic DNA from F1 pups using U3-H1 primers. The combination of U3 forward and H1 reverse amplifies the H1 promoter portion of the H1-siGFP cassette. The positive F1-36 pup is shown in red. Persistence of the GFP allele was confirmed by PCR using GFP primers. (d) Shining with fluorescence lamp of F1 littermate pups at day 3 of age (pictures were taken with a digital camera). (e) Embryonic day-13 embryos were harvested from pregnant female F0-4, and GFP sequences were detected using GFP-specific PCR primers. F1 embryos positive for integrated copies of the siGFP cassette using U3-H1 primers are shown. The F1-6 embryo positive for both GFP and siGFP is shown in red. (f) Protein extracts from embryonic day-13 embryos were analyzed by probing with a GFP antibody. Shown are embryos 1, 6, and 7. Reprobing with β-actin antibody was used as a loading control.