Isolation and expression of zebrafish zinc-finger transcription factor gene tsh1
Section snippets
Results
The Drosophila transcription factor teashirt (tsh) is involved in trunk segmentation, midgut morphogenesis and proximal leg formation (Alexandre et al., 1996, de Zulueta et al., 1994, Erkner et al., 1999, Mathies et al., 1994, Roder et al., 1992). Tsh functions in parallel with other homeotic proteins such as those encoded by the HOM-C complex genes (de Zulueta et al., 1994). In addition, tsh is required for transcriptional repression mediated by the Wingless signaling pathway (Gallet et al.,
Zebrafish husbandry
Wild-type zebrafish AB strains and neckless (nlsi26) (Begemann et al., 2001) were raised at 28.5 °C according to (Westerfield, 2000). Embryos were produced by pair-mating and staged as described (Kimmel et al., 1995).
RNA isolation and cDNA subtraction
Total RNA was isolated using TRIZOL RNA isolation kit (Gibco). mRNA was purified from total RNA using PolyA Tract-1000 system (Promega). cDNA subtraction was performed using Repressive Subtractive Hybridization Kit (Clontech).
Amplification of 5′ end of tsh1
The 5′ end of tsh1 was amplified by PCR using Smart RACE
Acknowledgements
We thank Lucille Joly for technical assistance and Dr. Marie-Andrée Akimenko for helpful discussions. The project was partly funded by Grant MOP-14460 from the Canadian Institutes of Health Research to M.E., Grants from NIH (1P20RR17703-04) and the Whitehall foundation (2002-12-103) to H.W., and the Cornell Vertebrate Functional Genomics Initiative fund to Q.L.
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Present address: Laboratory of Molecular Genetics, National Institute of Child Health and Development, National Institutes of Health, Bethesda, MD 20892, USA.