Identification of the lacZ insertion site and beta-galactosidase expression in transgenic chickens.
The quail:chick chimera system is a classical research model in developmental biology. An improvement over the quail:chick chimera system would be a line of transgenic chickens expressing a reporter gene. Transgenic chickens carrying lacZ and expressing bacterial beta-galactosidase have been generated, but complete characterization of the insertion event and characterization of beta-galactosidase expression have not previously been available. The genomic sequences flanking the retroviral insertion site have now been identified by using inverse polymerase chain reaction (PCR), homozygous individuals have been identified by using PCR-based genotyping, and beta-galactosidase expression has been evaluated by using Western analysis and histochemistry. Based upon the current draft of the chicken genome, the viral insertion carrying the lacZ gene has been located on chromosome 11 within the predicted gene for neurotactin/fractalkine (CX3CL1); neurotactin mRNA expression appears to be missing from the brain of homozygous individuals. When Generation 2 (G2) lacZ-positive individuals were inter-mated, they generated 361 G3 progeny; 82 were homozyous for lacZ (22.7%), 97 were wild-type non-transgenic (26.9%), and 182 (50.4%) were hemizygous for lacZ. Western analysis revealed the highest expression in the muscle and liver. With the identification of homozygous birds, the line of chickens is now designated NCSU-Blue1.
Mozdziak
Paul E
PE
Department of Poultry Science, North Carolina State University, Raleigh, NC 27695, USA. pemozdzi@unity.ncsu.edu
Wu
Qian
Q
Bradford
Jennifer M
JM
Pardue
Samuel L
SL
Borwornpinyo
Suparerk
S
Giamario
Carol
C
Petitte
James N
JN
D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D015139:Blotting, Southern; D054428:Chemokine CX3CL1; D020523:Chemokines, CX3C / Q000378:metabolism; D002642:Chick Embryo; D002645:Chickens; D005260:Female; D005838:Genotype; D007763:Lac Operon; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D012689:Sequence Homology, Nucleic Acid; D001616:beta-Galactosidase / Q000378:metabolism*
D054428:Chemokine CX3CL1; D020523:Chemokines, CX3C; D008565:Membrane Proteins; D001616:beta-Galactosidase