The cloning and characterization of chick tyrosinase from a novel embryonic cDNA library.
Very little is known about the genes involved in the regulation of avian skin and feather pigmentation. In mammals, two gene families have been identified as being important for the regulation of melanin biosynthesis. To isolate the avian equivalents of these families, we have generated an embryonic chick melanocyte cDNA library. Neural crest cells from 500 black chick embryos were cultured under conditions supportive of melanocyte differentiation and proliferation. A cDNA library was constructed and screened with a mouse tyrosinase cDNA probe. Nineteen clones were obtained, seven of which cross-hybridized to a mouse tyrosinase cDNA on Southern blots. The longest of these clones, B8.3 (1.9 kb), was sequenced and found to share 99.7% nucleotide and 99.8% amino acid sequence homology to a reported chick tyrosinase cDNA. Both Northern blot analysis and in situ hybridization demonstrated that clone B8.3 was expressed in the retinal pigment epithelium of chick embryos. Our results suggest therefore that the cDNA library described here may allow the cloning of novel melanogenic genes.
April
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Department of Anatomy and Cell Biology, Medical School, University of Cape Town, South Africa.
Franz
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Kidson
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D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002478:Cells, Cultured / Q000201:enzymology / Q000502:physiology; D002642:Chick Embryo; D003001:Cloning, Molecular; D015342:DNA Probes / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004815:Epidermal Growth Factor / Q000737:chemistry / Q000235:genetics; D005241:Feathers / Q000166:cytology / Q000502:physiology*; D018507:Gene Expression Regulation, Developmental / Q000502:physiology; D015723:Gene Library; D008544:Melanocytes / Q000201:enzymology / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D014442:Monophenol Monooxygenase / Q000235:genetics* / Q000378:metabolism; D010857:Pigment Epithelium of Eye / Q000201:enzymology / Q000502:physiology; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000032:analysis; D015183:Restriction Mapping; D012867:Skin / Q000166:cytology; D012879:Skin Physiological Phenomena
D015342:DNA Probes; D018076:DNA, Complementary; D012333:RNA, Messenger; D004815:Epidermal Growth Factor; D014442:Monophenol Monooxygenase