Genomic organization of the gene for pig zona pellucida glycoprotein ZP1 and its expression in mammalian cells.
The pig zona pellucida (pZP) is composed of glycoproteins pZP1, pZP3 alpha and pZP3 beta. Previously, we showed that pZP1 was proteolytically cleaved into two components (pZP4 and pZP2), and suggested that pZP1 was a homologue of mouse ZP2. We now report the genomic organization of pZP1 gene, and expression of the recombinant protein in mammalian cells. Two genomic clones, pZP4g and pZP4.1, were isolated by screening the pig genomic library with an already cloned cDNA encoding pZP4 as a probe. These clones covered 25 kb of the genome, including the entire pZP1 locus. The pig ZP1 genome consists of 18 exons and 17 introns. For production of the recombinant proteins of pZP1, a minigene containing a genomic fragment covering exon 6 to exon 17 was constructed and cloned into the expression vector, pBCMGS-neo. Mammalian cell lines, CHO (for stable expression) and 293T (for transient expression), transfected with the plasmid construct synthesized pZP1 recombinant proteins, probably as a membrane-bound protein.
Yamasaki
N
N
Department of Obstetrics and Gynecology, Hyogo College of Medicine, Japan.
Tsubamoto
H
H
Hasegawa
A
A
Inoue
M
M
Koyama
K
K
D000818:Animals; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D004527:Egg Proteins / Q000235:genetics*; D005260:Female; D015698:Genomic Library; D008322:Mammals / Q000235:genetics*; D008562:Membrane Glycoproteins / Q000235:genetics*; D011956:Receptors, Cell Surface / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis*; D014162:Transfection; D015044:Zona Pellucida / Q000502:physiology*; D000072216:Zona Pellucida Glycoproteins
D004527:Egg Proteins; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D000072216:Zona Pellucida Glycoproteins; C000606519:Zp1 protein, mouse; C000606520:Zp2 protein, mouse