title: Analyzing the promoters of two CYP9A genes in the silkworm Bombyx mori by dual-luciferase reporter assay.
abstract: Cytochrome P450s (CYPs) are widespread proteins that interact with exogenous chemicals from the diet or the environment. CYP9A subfamily genes are important in the silkworm Bombyx                 mori. We previously reported transcriptional levels of two CYP9A genes in different tissues and their responses to sodium fluoride (NaF). In this study, promoter truncation analysis using a dual-luciferase reporter assay in B. mori ovary cells (BmN) showed that the regions -1,496 to -1,102 bp for CYP9A19, and -1,630 to -1,210 bp for CYP9A22 were essential for basal transcriptional activity. Sequence analysis of these regions revealed several transcriptional regulatory elements but no typical promoter elements. Promoter activities were regulated after NaF induction and with an obvious dose effect. Although the dual-luciferase assay has been widely used to determine the activity of a given promoter in cell lines, problems with it still exist. Our results indicate that both plasmid size and construct protocols affect the experimental results.
authors.affiliation: School of Biology and Basic Medical Sciences, Soochow University, No. 199 Ren'ai Road, Dushu Lake Higher Education Town, Suzhou Industrial Park, Suzhou, 215123, Jiangsu, People's Republic of China.
authors.forename: Si-Si
authors.initials: SS
authors.lastname: Zhao
authors.forename: Guo-Dong
authors.initials: GD
authors.lastname: Zhao
authors.forename: Tian-Yuan
authors.initials: TY
authors.lastname: Di
authors.forename: Hua
authors.initials: H
authors.lastname: Ding
authors.forename: Xiao-Ling
authors.initials: XL
authors.lastname: Wan
authors.forename: Bing
authors.initials: B
authors.lastname: Li
authors.forename: Yu-Hua
authors.initials: YH
authors.lastname: Chen
authors.forename: Ya-Xiang
authors.initials: YX
authors.lastname: Xu
authors.forename: Wei-De
authors.initials: WD
authors.lastname: Shen
authors.forename: Zheng-Guo
authors.initials: ZG
authors.lastname: Wei
mesh_terms: D000818:Animals; D001483:Base Sequence; D012831:Bombyx / Q000201:enzymology* / Q000235:genetics; D002460:Cell Line; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic; D055785:Gene Knockdown Techniques; D017930:Genes, Reporter; D019476:Insect Proteins / Q000235:genetics*; D049409:Luciferases, Firefly / Q000096:biosynthesis / Q000235:genetics; D049410:Luciferases, Renilla / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D034741:RNA, Small Interfering / Q000235:genetics; D017422:Sequence Analysis, DNA; D012969:Sodium Fluoride / Q000494:pharmacology; D014162:Transfection
chemical_list: D019476:Insect Proteins; D034741:RNA, Small Interfering; D012969:Sodium Fluoride; D003577:Cytochrome P-450 Enzyme System; D049410:Luciferases, Renilla; D049409:Luciferases, Firefly