RT-PCR assay MYCT-1 gene in GES-1 cells.doc

 PAGE \* MERGEFORMAT 7 RT-PCR assay MYCT-1 gene in GES-1 cells [Abstract] Objective MYCT-1 (myc target) gene in gastric mucosa cell line GES-1 expression of GES-1 cells as a cell model of RNA interference. Methods extracted using TRIZOL GES-1 cell total RNA, RT-PCR amplification of MYCT-1 cDNA, DNA sequencing. results in GES-1 cells detected 504bp MYCT-1 cDNA fragments, sequencing confirmed with GenBank reported sequences. Conclusion GES-1 cells in the presence MYCT-1 expression, RNA interference can be used as cell model. [Keywords:] MYCT-1; GES-1 cell line; RT-PCR MYCT-1 (myc target 1) was cloned in our laboratory a new candidate tumor suppressor gene, GenBank accession number NM_025107 [1], has been named MTLC (myc target from laryngeal cancer cells). Preliminary study showed that MYCT-1 is widely expressed in a variety of normal tissue, and in gastric cancer, laryngeal cancer, liver cancer, breast cancer and other malignant tissues down [2 5]. MYCT-1 for further study the biological function of genes in cancer development and the role of , ready to use RNA interference (RNA interence, RNAi) technology to silence the normal cells MYCT-1 expression, in order to understand MYCT-1 low expression of normal cell growth, cell proliferation, cell cycle and apoptosis. In this paper, RT -PCR method and DNA sequencing to detect GES-1 cells in the gastric mucosa MYCT-1 expression of GES-1 cells were used as RNA interference model of value. 1 Materials and methods 1.1 Materials GES-1 cells (Cancer Institute); TRIZOL reagent, 1640, fetal bovine serum (FBS) (Gibco Company); reverse transcriptase, Taq enzyme (Promega Corporation); MYCT-1 primers (Takara Company ), the sequence was: forward primer: 5’-ATGGATCCCTGCACTGGCTGATGAGTGTGTA-3 ‘; reverse primer: 5’-GTAAGCTTGAACAGTGCCTTCACCCTCGA GGT-3’; other reagents were of analytical grade. 1.2 cell culture to 1.5 105 GES-1 cells were inoculated to 35mm Petri dish, containing 10% FBS 1640 medium, in 5% CO2, 37