调控区研究方法.doc

Experimental procedures to identify cis-acting elements and their DNA-binding proteinsPlant systems for the analysis of cis-acting elementsTransgenic plants.Transient expression using microprojectiles.Transient expression using protoplasts.Experimental procedures for the determination of cis-acting elementsDeletion analysis of promoter regions (loss-of-functionexperiment).Functional analysis of promoter fragments (gain-of-functionexperiment).cis-Element analysis using point mutations in promoter fragments(base substitution).Chromatin immunoprecipitation (ChIP) 48.48 He, Y. et al. (2003) Regulation of flowering time by histone acetylationin Arabidopsis. Science 302, 17511754Identification and confirmation of DNA-binding proteins for cis-acting elementsGel mobility shift assay.DNA footprint analysis.Methylation interference.DNA affinity precipitation (DNAP) 49.49 Phares, W. et al. (1992) The kappa B enhancer motifs in humanimmunodeficiency virus type 1 and simian virus 40 recognize differentbinding activities in human Jurkat and H9 T cells: evidence for NFkappaB-independent activation of the kappa B motif. J. Virol. 66,74907498Isolation of cDNA clones of the DNA-binding proteins forcis-acting elementsSouth-western blotting screening 20.20 Niu, X. et al. (2002) Maize ABI4 binds coupling element1 in abscisicacid and sugar response genes. Plant Cell 14, 25652576Yeast one-hybrid screening 29.29 Stockinger, E.J. et al. (1997) Arabidopsis thaliana CBF1 encodes anAP2 domain-containing transcription activator that binds to theC-repeat/DRE, a cis-acting DNA regulatory element that stimulatestranscription in response to low temperature and water deficit. Proc.Natl. Acad. Sci. U. S. A. 94, 10351040Candidate selection based on genomic data, such as phylogeneticanalysis and microarray analysis 50.50 Satoh, R. et al. (2004) A novel subgroup of bZIP proteins functions astranscriptional activators in