Figure 1.
Maps of the pTGZC (A) and pZCpTG (B) vectors.
Figure 2.
Co-expression of the four fluorescent proteins in PFFs and blastocysts.
(A) PFFs transfected with the linearized pZCpTG vector were observed under a confocal microscope using appropriate filters. The scale bar represents 10 µm. (B) Several reconstructed embryos derived from the pZCpTG PFFs were cultured until they reached the blastocyst stage. The blastocysts were observed under a fluorescence microscope using appropriate filters.
Table 1.
Development ability of nuclear transfer embryos derived from pZCpTG-PFFs and WT-PFFs.
Figure 3.
Genotype identification of multi-transgenic piglets.
(A) Positions of the two primer pairs for genotype identification. Genomic DNA from Piglet 2, 3, 4, 6, 7, 9, 10 was extracted, whereas ZC (B) and TG (C) bicistronic cassettes in the genome were detected by PCR using appropriate primers. Lane 1, DNA marker; lane 2, positive control; lane 3, negative control; lane 4–10, genomic DNA from Piglet 2, 3, 4, 6, 7, 9 and 10.
Figure 4.
Co-expression of the four fluorescent proteins in the fibroblast isolated from multi-transgenic piglets.
(A) Fibroblasts isolated from the ear tissue of a representative animal (Piglet 10) observed under a confocal microscope using appropriate filters. The scale bar represents 10 µm. (B) The fluorescence intensities of the four fluorescent proteins were measured. The results are shown as mean ± S.D.
Figure 5.
Co-expression of the four fluorescent proteins in various tissues of a multi-transgenic piglet.
Liver, kidney, hoof, heart, tongue, skin, nose and lung tissues from Piglet 7 were cryosectioned and observed under a confocal microscope using appropriate filters. The scale bar represents 100 µm.
Figure 6.
Expression profile analysis of four fluorescent proteins in various tissues by RT-PCR.
Total mRNA was extracted from various tissues of Piglet 7. The mRNA levels of the target genes (tdTomato, EGFP+ECFP and ZsYellow1) were determined by RT-PCR. The results are shown as mean ± S.D.
Figure 7.
Fluorescence in living multi-transgenic piglets.
Florescence on the noses and hooves of multi-transgenic piglets was directly observed using goggles. (A) Bright light. (B) tdTomato fluorescence. (C) EGFP fluorescence. (D) ECFP fluorescence. A WT piglet was used as negative control.