Novel double promoter approach for identification of transgenic animals: A tool for in vivo analysis of gene function and development of gene-based therapies.
Advances in vertebrate genetics have allowed studies of gene function in developing animals through gene knockout and transgenic analyses. These advances have encouraged the development of gene-based therapies through introduction of exogenous genes to enhance and/or replace dysfunctional or missing genes. However, in vertebrates, such analyses often involve tedious screening for transgenic animals, such as PCR-based genotype determinations. Here, we report the use of double-promoter plasmids carrying the transgene of interest and the crystallin-promotor-driven Green fluorescent protein (GFP) in transgenic Xenopus laevis tadpoles. This strategy allows a simple examination for the presence of GFP in the eyes to identify transgenic animals. PCR-based genotyping and functional characterization confirms that all animals expressing GFP in the eyes indeed carry the desired promoter/transgene units. Thus, the use of this and other similar vectors should dramatically improve current transgenesis protocols and reduce the time and cost for identifying transgenic animals.
Unit on Molecular Morphogenesis, Laboratory of Gene Regulation and Development, NICHD, NIH, Bethesda, Maryland 20892, USA.
Liezhen
L
Fu
Daniel
D
Buchholz
Yun-Bo
YB
Shi
D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D005821:Genetic Techniques; D015316:Genetic Therapy; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D007814:Larva / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics; D020782:Matrix Metalloproteinases / Q000235:genetics; D011401:Promoter Regions, Genetic; D019076:Transgenes; D014982:Xenopus laevis; D038222:gamma-Crystallins / Q000235:genetics
D008164:Luminescent Proteins; D038222:gamma-Crystallins; D049452:Green Fluorescent Proteins; D020782:Matrix Metalloproteinases