This transgenic medaka was established to rescue the fish from the fused centrum (fsc) phenotype by using an 8.4 kb genomic fragment encompassing the wnt4b gene as a transgene. The transgene has a 3.2 kb wnt4b promoter region that contained the cis-regulatory elements necessary for expression in the floor plate but not in the otic vesicles. In addition, a reporter gene that expressed EGFP under the control of a zebrafish alpha A-crystallin promoter is inserted into the 3'side of the 8.4 kb transgene.
The transgenic specimens have the exogenous wnt4b gene on a homozygous fsc genomic background. The exogenous wnt4b functions in the floor plate of the transgenic embryos and rescues these transgenic specimens from the fsc phenotype. The transgenic specimens are distinguish from other siblings easily by observing EGFP expression in their lens under a fluorescence stereomicroscope.
The mutaion of fsc is identical to that of fused mutant (MT51 and MT1122).
|Special affairs [Address]||
■ The RECIPIENT shall refer the following publication in any PUBLICATIONS.
(Name of the Publication: Inohaya K, Takano Y, Kudo A. (2010). Production of Wnt4b by floor plate cells is essential for the segmental patterning of the vertebral column in medaka. Development. 137, 1807-1813.)
|Organization||National Institute for Basic Biology|
|Frozen sperm in NIBB||○|
|Deposited by||Tokyo Institute of Technology|
|Document (PDF)||Genotyping protocol_wnt4b^fsc|
(A, B) Merged images of EGFP (green) expression and ALC (red) staining. Lateral views of the hatched larvae. Anterior to the left. The transgenic rescue larva shows normal segmental pattern in their vertebral column (A), whereas their sibling shows the fused centra and the defective intervertebral ligaments (B). Arrow head in A indicates the EGFP-positive lens in the transgenic rescue specimen.
A 8.4kb genomic fragment encompassing the wnt4b gene from the genomic DNA of the medaka Qurt line was cloned into the pGEM-T Easy vector (Promega). In addition, a reporter gene, which expresses EGFP under the control of a zebrafish alpha A-crystallin promoter, was subcloned into the SacI site on the 3' side of the 8.4 kb transgene.