AUTOMATED MICROSATELLITE GENOTYPING

DESCRIPTION: (Adapted from the Applicant's Abstract). Dense genetic linkage maps enable the identification of causative genes for genetic disease. Using these maps to genetically localize even one disorder on the genome requires hundreds of thousands of genotyping experiments. This genotyping process has been automated through the use of PCR-based microsatellite markers, automated DNA sequencers, robotic handling, and computer analysis. However, the microsatellite markers produce an intrinsic PCR stutter artifact that has thus far precluded full genotyping automation. Recently computational methods have been developed that overcome this key molecular biology bottleneck. The applicants software system automatically transforms DNA sequencer data files into genotypes. Limited empirical testing of such software system on DNA sequencer data has produced genotypes at a low error rate. These initial results suggest that further testing and validation may establish the feasibility of this computational approach. If ultimately successful, the methods described may help advance the accurate and rapid localization of genes within the human genome. This application focuses on determining the feasibility of (1) automated microsatellite genotyping with low error on data from automated DNA sequencers and (2) automated microsatellite genotyping using pooled DNA material. $ = TOTAL AWARD AMTS & NOT LIMITED TO PORTION OF PROJECT RELATED TO SUBJECT OF SEARCH SUBPROJECT $ = TOTAL AWARD AMOUNT DIVIDED BY NUMBER OF SUBPROJECTS SOURCE: CRISP FORMAT F FY 97 LAST UPDATE 04-07-98 1QUERY 1536 ID SEARCH 06/01/98 PAGE 390 --PROJECT NUMBER......1 R43 HG01589-01A1 INVESTIGATOR NAME/ADDRESS FY 97 SCHMUKLER, ROBERT E IRG/INTRAMURAL UNIT..ZRG7 PORE SQUARED BIOENGINEERING AWARD AMOUNT......... $100,000 13905 VISTA DR ROCKVILLE, MD 20853-2612 PERFORMING ORGANIZATION: PORE SQUARED BIOENGINEERING TITLE LOW VOLTAGE ELECTROPORATION FOR GENE TRANSFER ABSTRACT: DESCRIPTION (Adapted from Applicant's Abstract): The objective of the proposed research is to evaluate the gene transfer efficiency of a new method of electroporating cells, using low voltage (< 20V) waveforms, under physiologic conditions. The new method relies upon the unique structure and geometry resulting from embedding living cells into the holes of thin (10 um) insulating films (track-etched filters). This preparation is analogous to a multi-parallel-whole-cell-patch-clamp preparation. A third of a million cells are simultaneously embedded into the film holes and electroporated. The resistive seal around each cell of between 65-70 MQ serves to amplify the field strength across the film, by 1000X. This method allows electroporation to be carried out, while minimizing deleterious effects from joule heating and electrochemical products. One advantage is that only a single side of the cell membrane is porated, due to the asymmetric cellular geometry of the embedded cells. The ability of this new method to produce electroporation with 95-99% cell viability has already been demonstrated in a pilot study, Gene transfer efficiency will be determined by transfecting the B- Galactosidase gene into mammalian COS-7 monkey cells. The specific aim of the proposed research is to determine the gene transfer efficiency for this new method. $ = TOTAL AWARD AMTS & NOT LIMITED TO PORTION OF PROJECT RELATED TO SUBJECT OF SEARCH SUBPROJECT $ = TOTAL AWARD AMOUNT DIVIDED BY NUMBER OF SUBPROJECTS SOURCE: CRISP FORMAT F FY 97 LAST UPDATE 04-07-98 1QUERY 1536 ID SEARCH 06/01/98 PAGE 391 --PROJECT NUMBER......1 R43 HG01597-01 INVESTIGATOR NAME/ADDRESS FY 97 SCHULTZ, SHELDON IRG/INTRAMURAL UNIT..ZRG2 SEASHELL TECHNOLOGY AWARD AMOUNT......... $99,750 9265 DOWDY DRIVE, SUITE 105 SAN DIEGO, CA 92126 PERFORMING ORGANIZATION: SEASHELL TECHNOLOGY, LLC TITLE NEW TECHNIQUE FOR HIGH RESOLUTION MAPPING OF DNA ABSTRACT: DESCRIPTION: (Adapted from the Applicant's Abstract). The major goal of this project is to develop a new reliable, high resolution, efficient, and cost effective technique for the general detection of genetic defects in the human genome, and in selected pieces of DNA. The approach is to utilize the extraordinary capabilities of plasmon resonant particles which can provide non-toxic, very bright, robust, color-labeled probes that can be bound to any desired specific-locus DNA fragment, and then hybridized to the DNA strand of interest. The investigators expect to achieve a spatial resolution between marker probes of less than 0.5 KB. The investigators shall develop the specific reagents, protocols, and associated technology which will be user- friendly, and affordable for wide scale clinical applications. $ = TOTAL AWARD AMTS & NOT LIMITED TO PORTION OF PROJECT RELATED TO SUBJECT OF SEARCH SUBPROJECT $ = TOTAL AWARD AMOUNT DIVIDED BY NUMBER OF SUBPROJECTS SOURCE: CRISP FORMAT F FY 97 LAST UPDATE 04-07-98 1QUERY 1536 ID SEARCH 06/01/98 PAGE 392 --PROJECT NUMBER......1 R43 HG01664-01 INVESTIGATOR NAME/ADDRESS FY 97 SPITZNER, JOSEPH IRG/INTRAMURAL UNIT..ZRG2 TOPOGEN, INC AWARD AMOUNT......... $99,800 COLUMBUS, OH 43212-1155 PERFORMING ORGANIZATION: TOPOGEN, INC. TITLE GRAPHIC INTERFACE AND PROTOCOLS FOR GENOMIC DISPLAYS ABSTRACT: A growing need for graphically displaying and accessing genetic sequence information has been generated by the large (and increasingly larger) quantity of sequence data available to researchers. Despite this need, there are no generally acceptable standards for encoding and transmitting graphic display parameters (i.e., how to display a sequence and which features to highlight). MapKit (TopoGEN software) has been rated the best sequence mapping software of 1996 by Biotechnology Software and Internet Journal. On the Windows/PC platform, MapKit accomplishes the goals of graphically representing a broad range of sequence features and providing a local server that can interact with other applications to display and select sequence features. The purpose of this SBIR is to enable the development of a public domain standard that supports distributed data processing and is platform independent, i.e., a graphic sequence feature representation protocol that may be transmitted over the Internet and implemented on any computer system (Macintosh, Unix-based, etc.). The specific aims of the SBIR include developing the protocol, defining display and access standards for sequence viewers, creating as a prototype a public domain sequence viewer for the Windows/PC platform, creating a web site to support the project, and using the web site to distribute the protocol description and Windows software to users (genetics researchers, molecular biologists, etc.), software developers, and database managers.