The interaction of high-resolution electrophoresis and computational analysis in genome mapping
The construction of physical maps and the determination of the DNA sequence of chromosome-size segments of the human genome is a complex, multidisciplinary undertaking. The approach we have taken to construct a physical map and sequence of human chromosome 19 typifies these interactions. We exploit the power of both acrylamide and agarose gel electrophoresis to provide a simple and versatile method for DNA fingerprinting and the creation of contigs or sets of overlapping genomic clones. Cosmid libraries are constructed from Yeast Artificial Chromosomes (YAC) clones or from flow-sorted chromosomes. Cosmid DNA isolated from the screened library array is cut with a combination of five restriction enzymes and the fragment ends labeled with one of four different fluorochromes. Our approach to contig construction uses a robotic system to label restriction fragments from cosmids with fluorochromes, use of an automated DNA sequencer to capture fragment mobility data in a high resolution multiplex mode processes the mobility data to determine fragment length and provide a statistical measure of overlap among cosmids; and display the contigs and underlying cosmids for operator interaction and access to a database. Data analyses and interactions are conducted over a network of SUN workstations using a set of software tools that we developed and coupled to a commercially available database. Applying these methods, we have analyzed 5154 cosmid clones and assembled 515 contigs for chromosome 19. Some of these contigs have been identified with known genes and many have been mapped to the chromosome by fluorescence in situ hybridization. Existing contigs are being extended by a combination of walking and fingerprinting. 21 refs., 2 figs.
- Research Organization:
- Lawrence Livermore National Lab., CA (USA)
- Sponsoring Organization:
- DOE/ER
- DOE Contract Number:
- W-7405-ENG-48
- OSTI ID:
- 6698612
- Report Number(s):
- UCRL-102887; CONF-9004221-3; ON: DE90015363
- Resource Relation:
- Conference: 1. international conference on electrophoresis, superconducting and the human genome, Tallahassee, FL (USA), 10-13 Apr 1990
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
HUMAN CHROMOSOME 19
GENETIC MAPPING
ACRYLAMIDE
CHROMOSOMES
COSMIDS
DATA ANALYSIS
DNA SEQUENCING
DNA-CLONING
ELECTROPHORESIS
GELS
YEASTS
AMIDES
CLONING
COLLOIDS
DISPERSIONS
DNA HYBRIDIZATION
EUMYCOTA
FUNGI
HYBRIDIZATION
MAPPING
MICROORGANISMS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PLANTS
STRUCTURAL CHEMICAL ANALYSIS
550400* - Genetics
550200 - Biochemistry