IMPROVED RFLP ANALYSIS FOR GENETIC LINKAGE MAPPING

ONE OF THE MOST ACTIVE AND CHALLENGING FIELDS IN MOLECULAR BIOLOGY TODAY IS THE GENETIC STUDY OF EUKARYOTIC SYSTEMS. ONE RAPIDLY GROWING AREA IN THIS FIELD IS THE STUDY OF RESTRICTION FRAGMENT LENGTH POLYMORPHISMS (RFLPS). THIS PROCEDURE USES RADIOLABELED HYBRIDIZATION TECHNIQUES AND AUTORADIOGRAPHY TO CONSTRUCT A HUMAN GENETIC LINKAGE MAP BY IDENTIFYING GENETIC MARKERS ALONG THE GENOME. 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 QUANTITATION OF THE RADIOACTIVE BANDS BECAUSE OF THE LIMITED DYNAMIC RANGE OF FILM AND THE RESOLUTION PROBLEMS WHICH ARISE WHEN PHOSPHOROUS-32 PROBES ARE USED. 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 A COMPUTER MONITOR FOR VISUALIZATION AND FURTHER DATA ANALYSIS. BY THIS APPROACH, THE RESEARCHER WILL BE ABLE TO AVOID THE DELAYS ASSOCIATED WITH AUTORADIOGRAPHY, AND 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.