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.
Fu
Liezhen
L
Unit on Molecular Morphogenesis, Laboratory of Gene Regulation and Development, NICHD, NIH, Bethesda, Maryland 20892, USA.
Buchholz
Daniel
D
Shi
Yun-Bo
YB
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