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Volume 129, Issue 2, Pages (August 2005)

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1 Volume 129, Issue 2, Pages 565-576 (August 2005)
Identification of Functional Genetic Variants in Cyclooxygenase-2 and Their Association With Risk of Esophageal Cancer  Xuemei Zhang, Xiaoping Miao, Wen Tan, Baitang Ning, Zhihua Liu, Yuan Hong, Wenguang Song, Yongli Guo, Xinyu Zhang, Yan Shen, Boqin Qiang, Fred F. Kadlubar, Dongxin Lin  Gastroenterology  Volume 129, Issue 2, Pages (August 2005) DOI: /j.gastro Copyright © 2005 American Gastroenterological Association Terms and Conditions

2 Figure 1 Sequence analyses of the COX-2 promoter region with an ABI PRISM 3700 automatic sequencer reveal 3 SNPs located at nucleotides (A) −1290 (A→G), (B) −1195 (G→A), and (C) −765 (G→C). The arrows localize the base changes at the nucleotide positions. Gastroenterology  , DOI: ( /j.gastro ) Copyright © 2005 American Gastroenterological Association Terms and Conditions

3 Figure 2 Transient reporter gene expression assays with constructs containing full-length COX-2 promoter. (A) Schematic of reporter gene constructs having a full-length COX-2 promoter, with the only difference between the 8 constructs being an A→G at the −1290, a G→A at −1195, and a G→C at −765 polymorphic sites. (B) Luciferase expression of the 8 constructs in HeLa cells cotransfected with pRL-SV40 to standardize transfection efficiency. Luciferase levels of pGL-3 Basic and pRL-SV40 were determined in triplicate and standardized for transfection efficiency. Fold increase was measured by defining the activity of the empty pGL-3 Basic vector as 1. Data shown are the means fold increase ± SD from 3 independent transfection experiments, each performed in triplicate. **P < .001 compared with each of the construct counterparts. Gastroenterology  , DOI: ( /j.gastro ) Copyright © 2005 American Gastroenterological Association Terms and Conditions

4 Figure 3 (A) Electrophoretic mobility shift assays with biotin-labeled oligonucleotides containing the −1195A or −1195G allele and nuclear extracts from HeLa cells. Lanes 1 and 7 show the mobilities of the labeled oligonucleotides without nuclear extracts; lanes 2 and 8 show the mobilities of the labeled oligonucleotides with nuclear extracts in the absence of competitor; and lanes 3–6 and 9–12 show the mobilities of the labeled oligonucleotides with nuclear extracts in the presence of various unlabeled competitors as indicated below the bottom autoradiograph. The arrow localizes a major oligonucleotide/nuclear protein complex. (B) Chromatin immunoprecipitation assays using antibodies against either c-MYB or β-actin. The presence of the COX-2 promoter was assayed for using PCR. The PCR products are shown here after electrophoreses on a 2% agarose gel. Gastroenterology  , DOI: ( /j.gastro ) Copyright © 2005 American Gastroenterological Association Terms and Conditions


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