[Construction of eukaryotic expression clone for human amelogenin].
OBJECTIVE
To construct-the eukaryotic expression clone for human amelogenin.


METHODS
Total RNA was isolated from human fetal tooth buds. RT-PCR was used to amplify the amelogenin encoding region, and the amplified fragment for human amelogenin was inserted into eukaryotic expression vector PcDNA 3.1. The positive clones were selected and analyzed by restriction endonuclease mapping and DNA sequencing.


RESULTS
570 bp fragment was produced by RT-PCR; it was of the same size as expected based on human ameloginin mRNA encoding area length. The sequence of the inserted fragment from the recombinant clone PcDNA 3.1-AMG was consistent with that of AMELX from GenBank with one mismatch on 485 from G to C, without affecting the amino acid sequence.


CONCLUSIONS
The eukaryotic expression clone PcDNA 3.1-AMG was successfully constructed with the properly inserted DNA sequence encoding mature human amelogenin.
Department of Periodontology, Guangdong Provincial Stomatological Hospital, Guangzhou 510280, China.
Jin-cai
JC
Zhang
Chun-yi
CY
Yin
Yun-hui
YH
Zhang
Chuan-jiang
CJ
Zhao
D053523:Amelogenin; D001483:Base Sequence; D002999:Clone Cells / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D004274:DNA, Recombinant / Q000096:biosynthesis / Q000235:genetics; D003746:Dental Enamel Proteins / Q000096:biosynthesis / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D005057:Eukaryotic Cells / Q000378:metabolism; D005333:Fetus; D005822:Genetic Vectors; D006801:Humans; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques; D017422:Sequence Analysis, DNA; D014083:Tooth Germ / Q000378:metabolism
C507199:AMELX protein, human; D053523:Amelogenin; D018076:DNA, Complementary; D004274:DNA, Recombinant; D003746:Dental Enamel Proteins