Cloning and embryonic expression of five distinct sfrp genes in the zebrafish Danio rerio
Section snippets
Phylogenic distribution of sFRPs
To date, eight distinct sFRPs have been identified, mostly in vertebrates (i.e., amphibians, birds, fishes and mammals), but also in the urochordate Ciona intestinalis (Dehal et al., 2002). One sFRP has been characterized in the sea urchin Strongylocentrotus purpuratus (Illies et al., 2002). Finally, no sFRP has been identified so far in protostome organisms, suggesting that sFRP arose after the separation between protostome and deuterostome lineages. In silico analyses of amino-acid sequences
Cloning of sfrp genes
Except for the gene sfrp3 (gift from M. Furutani-Seiki), the full-length cDNA sequence of sfrp1a, sfrp1b, sfrp2 and sfrp5 was obtained by PCR amplification using a cDNA library constructed using the Invitrogen cDNA single strand RT-PCR kit. Total RNA is extracted from 24 hpf zebrafish embryo using the Trizol reagent (Invitrogen). For the PCR amplification, oligonucleotides are synthesized using information obtained on sfrp gene sequences from the zebrafish genome database Ensembl available on
Acknowledgements
We are thankful to Dr. Jamile Hazan for the cloning of the sfrp1b gene. This study was supported by grants from the Wellcome trust and the Medical Research Council awarded to C.H.
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