Counting proteins bound to a single DNA molecule.
Chromosomes contain DNA covered with proteins performing functions such as architectural organization and transcriptional regulation. The ability to count the number of proteins bound to various regions of the genome is essential for understanding both architectural and regulatory functions. We present a straightforward method of counting gfp-conjugated proteins bound to an individual duplex DNA molecule by calibrating to a commercially available fluorescence standard using wide-field fluorescence microscopy. We demonstrate our method using the E. coli nucleoid-associated protein Fis.
Northwestern University, Department of Molecular Biosciences, 2205 Tech Drive, Hogan 2-100, Evanston, IL 60208-3500, United States. john-graham@northwestern.edu
John S
JS
Graham
Reid C
RC
Johnson
John F
JF
Marko
D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D035021:Factor For Inversion Stimulation Protein / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D008856:Microscopy, Fluorescence / Q000379:methods* / Q000592:standards
D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D035021:Factor For Inversion Stimulation Protein; C501034:Fis protein, E coli; D004247:DNA