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Different Expression of DOC-1 mRNA in Oral Squamous Cell Carcinoma and the Biological Influence of Exogenous p12DOC-1 on Tca8113 Cell

Author: RenJun
Tutor: SunMoYi；LiJianHu
School: Fourth Military Medical University
Course: Clinical Stomatology
Keywords: p12DOC-1 Transfection Oral squamous cell carcinoma Tca8113 cells DOC-1 RT-PCR
CLC: R739.8
Type: Master's thesis
Year: 2008
Downloads: 58
Quote: 0
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Abstract


Oral cancer is the most common tumors of the oral and maxillofacial region, located on the surface or near surface parts of its predilection, it seriously affected the patients with normal physiological function and social interaction. With the rapid development of molecular biology techniques, the occurrence of many diseases, the development has been obtained from the level of the gene corresponding explanation. Mutations of oncogenes and tumor suppressor genes, inactivation, abnormal expression of the cancer in recent years, a number of genes associated with oral cancer have been found occurrence of oral cancer research from the molecular level, the development may. Oral cancer gene to 1 (deleted in oral cancer-1, DOC-1) Harvard University Todd in 1995 was found in hamster oral epithelial a candidate tumor suppressor gene, p12 protein is the protein product of the gene encoding the study found that p12 The protein has the function of inhibiting tumor growth. Human DOC-1 is located on chromosome 12q24 cDNA full length of 1627bp encoding 115 amino acids, the protein product of the molecular weight of 12.4kDa. The purpose of our group preliminary study found that the different degree of differentiation of oral squamous cell carcinoma in p12 protein expression level differences, the purpose of the research is by semi-quantitative RT-PCR method to observe the DOC-1 gene in various degrees of differentiation mRNA expression levels in oral cancer is different. Building DOC-1 gene ORF coding region of eukaryotic expression vector, transfection of the the tongue cancer Tca8113 cell lines, to observe the behavior of the exogenous expression of p12 protein tongue cancer of Tca8113 cells in the biology. Method 1. Specific primers were designed using the RT-PCR method to detect oral squamous cell carcinoma and normal oral mucosa DOC-1 mRNA expression. After electrophoresis of the resulting product by computer gel imaging system, the gray value of each product was measured, the reference grayscale value and the inner by a relative value comparing each product, and then the relative values ??obtained were statistically analyzed. 2 designed for people DOC-1 gene ORF coding region primers, in HaCaT (immortalized human epidermal cells) cloned ORF coding region sequence, connected to the corresponding specific restriction sites at both ends of the sequence, by restriction enzyme digestion , ligation, transformation, sequencing, and successfully constructed the eukaryotic expression vector containing DOC-1 gene ORF coding region. Will build good eukaryotic expression vector containing DOC-1 gene ORF coding region by transfection kit cell lines transfected with tongue cancer Tca8113 stable transfected cell proliferative capacity and cell invasion ability was measured after transfection observed outside endogenous expression of p12 protein in the biological behavior of the cells of tongue cancer Tca8113. Results 1. DOC-1 mRNA in oral cancer grayscale value of 1.22 ± 0.12, gray-scale value of 1.58 ± 0.15 in the oral mucosa; DOC-1 mRNA in well-differentiated oral cancer gray value 1.35 ± 0.04, moderately differentiated oral cancer gray value of 1.20 ± 0.05, the gray in poorly differentiated oral cancer value of 1.08 ± 0.03. 2 build a good vector by double digestion, electrophoresis, DNA sequencing, show that containing DOC-1 gene ORF coding region of eukaryotic expression vector pEGFP-N1-DOC-1 was successfully constructed. Screened stable transfected cells and control cells compared reduced ability of cell proliferation, cell invasion and metastasis ability is also reduced, and compared with the control group cells, change with significant. Conclusion The present study demonstrated that the amount of DOC-1 mRNA expression differences in the degree of differentiation of oral squamous cell carcinoma, the lower degree of differentiation of oral squamous cell carcinoma, the lower the amount of DOC-1 mRNA expression. DOC-1 gene may play an important role in oral squamous cell carcinoma of the early low DOC-1 gene mRNA expression may be an early event in oral squamous cell carcinoma. Successfully constructed containing DOC-1 gene ORF coding region of the eukaryotic expression vector pEGFP-N1-DOC-1, Tca8113 cells transfected with tongue cancer been stably transfected stably transfected cells reduced proliferative capacity, cell invasion and metastasis also reduced , and compared with the control group cells, the change has a significant difference. The plasmid vector DNA stable transfection, and may be integrated with the host chromosome occurs by causing transfected cells change in biological traits.

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CLC: > Medicine, health > Oncology > Oral cavity, maxillofacial tumors
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