Toward understanding the function of amelogenin using transgenic mice.
The purpose of this study was to establish transgenic mouse lines as a tool to investigate the function of amelogenin during mineralization by causing ectopic production of amelogenin and studying its effect. The mouse amelogenin (mAme) was cloned from a 16-day-old whole mouse embryo cDNA library and was determined to be "full-length" mouse amelogenin (with a complete coding region) by comparison with the mouse amelogenin reported previously by Snead et al. (1985) and Lau et al. (1992). The overexpression construct contained: (1) the rat osteocalcin (OC) promoter (1.8 kb); (2) the adenovirus splicing casettes, including introgenic (Int) sequence (0.3 kb); (3) the full-length mAme cDNA (0.8 kb); and (4) the polyadenylation signal sequence from the pSG5 mammalian expression vector. Both Southern blotting and polymerase chain-reaction (PCR) analyses were performed, by means of a specific probe and a pair of oligodeoxynucleotides to OcIntmAme(A)+, respectively. The animals which showed transgene-positive in both analyses were further used to establish F1 animals. Heterozygocity was confirmed with F1 animals by PCR analysis of DNA from the F0 x FVB/N pups. Three independent transgenic F1 heterozygous lines (640t, 706t, and 708t) have now been established. The generation of F2 homozygous lines is under way. The heterozygous transgenic animals are currently being analyzed for alterations in the morphology and structure of various bone tissues.
Ibaraki-O'Connor
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Bone Research Branch, National Institute of Dental Research, National Institutes of Health, Bethesda, Maryland 20892, USA.
Nakata
K
K
Young
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MF
D053523:Amelogenin; D000818:Animals; D002417:Cattle; D002999:Clone Cells; D003001:Cloning, Molecular; D003746:Dental Enamel Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology*; D015870:Gene Expression; D005818:Genetic Engineering / Q000379:methods*; D051379:Mice; D008822:Mice, Transgenic / Q000235:genetics*; D015675:Osteocalcin / Q000235:genetics; D011401:Promoter Regions, Genetic; D051381:Rats; D017422:Sequence Analysis, DNA; D014074:Tooth Calcification / Q000502:physiology; D019076:Transgenes
D053523:Amelogenin; C507200:Amelx protein, mouse; C507201:Amelx protein, rat; D003746:Dental Enamel Proteins; D015675:Osteocalcin