Identification and analysis of regulatory proteins binding to the promoter of nicotine demethylase gene CYP82E4 in tobacco

To identify proteins regulating the expression of the nicotine demethylase gene CYP82E4,a promoter fragment of the gene was used as the bait for screening of a cDNA library using yeast one-hybrid system.The results showed that the yeast cotransformation efficiency of bait vector pHIS2/pE4 and prey vector pGADT7-Rec2 reached 1×106.Thirty-nine positive clones were obtained and sequenced with the insertion size between 850 to 2 500 bp.Among these clones,a promising zinc finger transcription factor which could be involved in regulation of target gene was identified and its function will be charaterized to provide information for development of low-nornicotine tobacco variety via molecular breeding.