PHOTOCHEMICAL MODIFICATION OF HYBRIDIZATION MEMBRANES

THE NUMBER OF NUCLEIC ACIDS TO MEMBRANE FILTERS HAS PROVIDED A NUMBER OF APPLICATIONS FOR MOLECULAR HYBRIDIZATION INCLUDING THE ANALYSIS OF GENOME STRUCTURE AND FUNCTION AND DIAGNOSIS OF INFECTIOUS DISEASES AND GENETIC DISORDERS. THE SENSITIVITY AND REPRODUCIBILITY OF THE HYBRIDIZATION REACTION DEPEND ON THE PROPERTIES OF THE MEMBRANE USED. BSI HAS DEVELOPED A PROPRIETARY PHOTOIMMOBILIZATION TECHNOLOGY THAT ALLOWS COVALENT COUPLING OF REAGENTS ONTO NUMEROUS MEMBRANE MATERIALS. WE WILL APPLY THIS TECHNOLOGY TO MODIFY THE SURFACE OF MEMBRANES USED IN NUCLEIC ACID HYBRIDIZATION ASSAYS. THE PHASE I OBJECTIVES ARE TO SYTHESIZE COATING REAGENTS WHICH WILL PERMIT THE ATTACHMENT OF PHOTOREACTIVE GROUPS TO MEMBRANE SURFACES THAT ARE CAPABLE OF COVALENTLY BONDING TO DNA; DEMONSTRATE THAT POSITIVELY CHARGED GROUPS USED TO IONICALLY BIND THE DNA CAN BE REMOVED AFTER DNA PHOTOIMMOBILIZATION IS COMPLETE TO REDUCE BACKGROUND PROBLEMS; AND DEFINE CONDITIONS FOR DNA TRANSFER, AND HYBRIDIZATION AND STRIPPING OF PROBES ON MODIFIED MEMBRANES. THE LONG-RANGE GOAL IS TO DEVELOP A MEMBRANE THAT CAN FIRMLY BIND DNA FRAGMENTS OF ALL SIZES, PROVIDE MAXIMUM SIGNAL WITH MINIMAL BACKGROUND, CAN BE REPROBED MULTIPLE TIMES, AND CAN BE USED IN A NUMBER OF FORMATS UNDER A WIDE RANGE OF CONDITIONS.