The purpose of this project is to develop new analytical methodology involving electrophores to speed up genetic mapping. An electrophore is a volatile compound which is detected in the gas phase by an electron capture technique. We have devised a means to use electrophores as labels for DNA probes. The most important property of electrophores for this purpose is their potential for extensive multiplicity. This refers to the opportunity to utilize many electrophores simultaneously as labels in a given assay. For example, each of many different DNA probes could be labeled with a unique electrophore, and then combined for use as a multi-probe cocktail. We propose to develop methodology for genetic mapping based on hybridization assays with electrophore-labeled DNA probes. The assays will be performed as dot blots in which each blot is hybridized with a solution containing many such probes. Many target sequences of DNA in the dot blot can thereby be tested simultaneously. In more detail, we propose in this startup project to establish a multiplicity of 12 for electrophore-labeled DNA probes. These probes will be combined and used to test do blots of Bluescript, lambda, phiX174 and adenovirus DNA. Detection will be accomplished by electron capture negative ion mass spectrometry (ECNI-MS) using a particle beam interface at a throughput of 15 seconds/sample. A gas chromatography (GC) interface will also be studied. Also presented are plans for future work, beyond the scope of this startup project, in which the multiplicity would be raised into the range of 100 to 400 while maintaining the same analysis speed of 15 seconds/dot blot. This would provide, for the detection step, a throughput of 24,000 hybridization assays per hour for a multiplicity of 100.
