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Abstract
7 E4 R' \( i3 H" hMultiple genetic modifications in pigs can essentially benefit research on agriculture, human disease and xenotransplantation.0 r3 _% {, l5 Z+ u
Most multi-transgenic pigs have been produced by complex and time-consuming breeding programs using multiple
4 x& l+ j: N6 {: ]7 isingle-transgenic pigs. This study explored the feasibility of producing multi-transgenic pigs using the viral 2A peptide in0 x) @4 Y& ]2 x8 f9 j1 N4 O
the light of previous research indicating that it can be utilized for multi-gene transfer in gene therapy and somatic cell, I a4 ~/ L: K9 I
reprogramming. A 2A peptide-based double-promoter expression vector that mediated the expression of four fluorescent) F- u" h) z( M% K
proteins was constructed and transfected into primary porcine fetal fibroblasts. Cell colonies (54.3%) formed under G4184 t S7 h R, ]' y" t8 [8 G
selection co-expressed the four fluorescent proteins at uniformly high levels. The reconstructed embryos, which were
% Y" d6 {$ _6 ~6 |/ F' D+ S7 Iobtained by somatic cell nuclear transfer and confirmed to express the four fluorescent proteins evenly, were transplanted
2 C m+ P. h' O* u& u4 p! binto seven recipient gilts. Eleven piglets were delivered by two gilts, and seven of them co-expressed the four fluorescent6 ?+ N! {6 n; [5 [
proteins at equivalently high levels in various tissues. The fluorescence intensities were directly observed at the nose, hoof
8 l1 y- ~. h, r7 Q$ Z0 i% A% e6 ~and tongue using goggles. The results suggest that the strategy of combining the 2A peptide and double promoters
g9 u! C" W& V) g# [7 cefficiently mediates the co-expression of the four fluorescent proteins in pigs and is hence a promising methodology to
- g+ [$ H5 L# k+ tgenerate multi-transgenic pigs by a single nuclear transfer. |
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发表于 2011-5-25 13:08
发表于 2011-5-25 13:14
