REAL TIME IMAGING SYSTEM FOR SCREENING DNA LIBRARIES

ONE OF THE MOST ACTIVE AND CHALLENGING FIELDS IN MOLECULAR BIOLOGY TODAY IS THE GENETIC STUDY OF EUKARYOTIC SYSTEMS. AN ESSENTIAL, BUT OFTEN RATE LIMITING STEP IN THIS AREA IS THE SCREENING OF DNA LIBRARIES FOR GENE MAPPING. THIS PROCEDURE USES RADIOLABEL HYBRIDIZATION TECHNIQUES AND AUTORADIOGRAPHY TO ACHIEVE THE REQUIRED SENSITIVITY AND SPECIFICITY. THE PARTICULAR PART OF THE PROCESS THAT THIS PROPOSAL ADDRESSES IS THE MEASUREMENT OF THE DISTRIBUTION OF RADIOACTIVITY ON THE TRANSFER MEMBRANE. NORMALLY THE AUTORADIOGRAPHS TAKEN OF THIS MEMBRANE REQUIRE AN EXPOSURE OF 16 TO 24 HOURS. FURTHERMORE, UNCERTAINTY IS INTRODUCED IN THE SELECTION OF THE CLONES OF INTEREST BECAUSE OF THE PROBLEMS OF ACHIEVING GOOD REGISTRATION BETWEEN THE TRANSFER MEMBRANE AND THE ORIGINAL AGAR DISH. WE PROPOSE TO UTILIZE A NEWLY AVAILABLE, POSITION SENSITIVE NUCLEAR DETECTOR TO GENERATE A REAL TIME IMAGE OF THE DISTRIBUTION OF THE RADIOACTIVITY ON THE TRANSFER MEMBRANE. FURTHERMORE, WE PROPOSE TO PROJECT THIS IMAGE DIRECTLY ONTO THE PRIMARY AGAR PLATE. BY THIS APPROACH, NOT ONLY WILL THE RESEARCHER WILL BE ABLE TO HARVEST THE APPROPRIATE PLAQUES FROM THE PLATE WITHOUT THE NEED TO SUFFER THE DELAYS ASSOCIATED WITH AUTORADIOGRAPHY, BUT WILL ALSO BE ABLE TO CARRY OUT THE PROCEDURE WITH A MUCH HIGHER LEVEL OF CONFIDENCE. SUCH AN ABILITY SHOULD SIGNIFICANTLY IMPROVE THE EFFICIENCY OF THE ENTIRE PROCEDURE AND ALLOW MORE RESOURCES TO BE DIRECTED TOWARD THE ANALYTICAL ASPECTS OF THE WORK.