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KMID : 0829220100340040179
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Korean Journal of Oral and Maxillofacial Pathology
2010 Volume.34 No. 4 p.179 ~ p.189
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Cloning and Identification of Novel Genes Expressed in Human Salivary Gland
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Kim Yeon-Sook
Lee Suk-Keun
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Abstract
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In order to unravel unidentified genes from human salivary gland, a cDNA library of human submandibular gland was constructed in the Uni-ZAP XR vector by use of mRNA from human submandibular gland and ZAP-cDNA® Gigapack® Ⅲ Gold Cloning Kit. cDNA of salivary gland was subtracted with cDNA of immortalized human keratinocyte cell line, Rhim Human Epithelial Keratinocyte cell line. The phage cDNA library was converted into a pBluescript phagemid cDNA library, which was subsequently plated on LB plates with ampicillin, IPTG, and X-gal, and white colonies were selected for sequencing. Among 200 clones analyzed, four clones containing C77-091, C75-014, C76-022, and C76-012 designated orphan genes that are intensely expressed in the interlobular ductal and serous acinar cells of human submandibular gland. Particularly C77-091 gene expresses 46 amino acids peptide (pI=9.45). C75-014 and C76-022 genes were characterized as those expressing excretory basic proteins primarily consist of alanine, proline, and leucine residues, mimicking a basic proline-rich protein (bPRP) showing helical structures and having multiple consensus sequences of phosphorylation sites. The strong expression of C76-012 mRNA in the nuclei of salivary ductal and acinar cells suggests a role of C76-012 gene as a DNA binding RNA/protein. These data suggest that the identification of four orphan genes from the human salivary glands may add further understanding of greater role of salivary proteins providing innate immunity by protecting and stabilizing the mucosal epithelium in the maintaining homeostasis of oral mucosa.
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KEYWORD
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Subtraction cloning, Human salivary gland, Oral mucosa, Novel genes
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