High throughput visualization and analysis of human short tandem repeat polymorphisms (STRP`s) using an infrared based automated DNA sequencer

Journal Article · Thu Sep 01 04:00:00 EDT 1994 · American Journal of Human Genetics

OSTI ID:134035

Jang, G Y; Gartside, B O; Brumbaugh, J A ^[1]

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1. LI-COR, Inc., Lincoln, NE (United States); and others

Short tandem repeat polymorphisms (STRPs) including di-, tri-, and tetranucleotide repeats are very useful markers for gene mapping, genetic diagnosis, and forensics because they are highly polymorphic, abundant throughout the mammalian genome, and amplifiable by the polymerase chain reaction (PCR). In order to meet the demand for large scale gene mapping and genetic diagnosis, the overall speed of genotyping STRPs should be increased. Automated detection systems using laser irradiation along with infrared fluorescently labeled PCR primers or dATP`s have been used for improved detection of PCR amplified STRPs when compared to conventional radioactive detection methods. Here, we report protocols that provide high throughput visualization and analysis of PCR amplified STRPs using the LI-COR Model 4000S DNA Sequencer. Short (15 cm separation distance) gels were used which produced rapid migration of the DNA fragments to the scanning detector (less than 1 hour from sample loading to detection of up to 350 base long DNA fragments). Seven percent denaturing acrylamide gels with a constant 2000V were employed for fast and adequate resolution. A 64 well format was used for loading (60 samples with 4 lanes for standard markers). Multiple loading of samples (using the same gel up to 3 times) has been achieved. In addition, we have multiplexed more than one locus per lane. By applying these conditions (60 samples x 3 loci x loads/gel x 2 gels/day) it is possible to generate images for over 1000 person loci in less than a day. Images were analyzed using Scanalytics RFLPscan software. The output data gave each allele size in number of base pairs.

• OSTI ID: 134035
• Report Number(s): CONF-941009--
• Journal Information: American Journal of Human Genetics, Journal Name: American Journal of Human Genetics Journal Issue: Suppl.3 Vol. 55; ISSN AJHGAG; ISSN 0002-9297
• Country of Publication: United States
• Language: English

Related Subjects

44 INSTRUMENTATION
INCLUDING NUCLEAR AND PARTICLE DETECTORS
55 BIOLOGY AND MEDICINE
BASIC STUDIES
AUTOMATION
BIOLOGICAL MARKERS
DETECTION
DNA SEQUENCERS
EFFICIENCY
FLUORESCENCE
GENES
GENETIC MAPPING
GENOTYPE
INFRARED SPECTROMETERS
MULTIPLEXERS
NUCLEOTIDES
POLYMERASE CHAIN REACTION
RESOLUTION
SIZE
STRUCTURAL CHEMICAL ANALYSIS