Gene inactivation by multiphoton-targeted photochemistry

Multiphoton-targeted photochemistry was used to selectively inactivate the expression of genes in vertebrate cells. A membrane permeable DNA-associati ng vital dye, ethidium bromide monoacetate (visible wavelength single photo n absorption peak at 530 nm) was used to photosensitize chromosomes in divi ding cells. A 100-ps infrared laser beam operating at 1.06 microns was focu sed onto a selected region of a mitotic chromosome corresponding to the sit es of the nucleolar (ribosomal) genes. Individual cells followed through mi tosis demonstrated a reduction in the number of nucleoli formed in daughter cells that corresponded to the number of nucleolar genes sites irradiated. These results demonstrate the ability to selectively manipulate genes by u sing the focal point specificity characteristic of multiphoton microscopy. This technique should have wide biotechnology applications both in vitro an d in vivo.