Efficient generation of functional transgenes by homologous recombination in murine zygotes.
To assess the feasibility of generating functional transgenes directly via homologous recombination between microinjected DNA fragments, three overlapping genomic DNA fragments, together constituting the human serum albumin (hSA) gene, were coinjected into murine zygotes. The resulting transgenic mice were analyzed for structure and expression of the transgene. All transgenic mice carried recombined hSA DNA fragments and 74% contained a reconstituted hSA gene. HSA expression could be detected in liver and serum in most (72%) of these animals. Only correctly sized hSA transcripts were observed. Transgenic hSA could not be distinguished from the human serum-derived protein by radioimmunoassay or Western blotting. The high frequency and accuracy of homologous recombination in murine zygotes reported here allows the efficient generation of relatively large transgenes.
Pieper
F R
FR
GenePharming Europe B.V., Leiden University, The Netherlands.
de Wit
I C
IC
Pronk
A C
AC
Kooiman
P M
PM
Strijker
R
R
Krimpenfort
P J
PJ
Nuyens
J H
JH
de Boer
H A
HA
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D015153:Blotting, Western; D004247:DNA; D004274:DNA, Recombinant; D015870:Gene Expression; D005821:Genetic Techniques; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011863:Radioimmunoassay; D011995:Recombination, Genetic; D012709:Serum Albumin / Q000235:genetics; D015053:Zygote / Q000378:metabolism*
D004274:DNA, Recombinant; D012709:Serum Albumin; D004247:DNA