Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Patent
1997-05-29
1998-12-08
Arthur, Lisa B.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
935 77, 935 78, 536 2532, 204451, 204601, 4352852, 4352872, C12M 100, C12M 304, C12N 1500, C07H 2100
Patent
active
058467270
ABSTRACT:
A system is disclosed for the rapid and cost-effective sequencing of DNA. There are three principal components of the system: (1) a microreactor, which prepares DNA sequencing "ladders" using solid-phase techniques, preferably in capillary tubes whose volumes are on the order of 10-1000 nanoliters, preferably 10-200 nanoliters; (2) a microfabricated electrophoresis capillary separation unit; and (3) a fluorescence detector with single-mode optical fibers interfaced directly to the electrophoresis capillary. The system is suitable for a highly multiplexed, automated DNA sequencing device Typical. steps in sequencing are as follows: (1) PCR amplification of a DNA template in microtiter dishes using labelled primers, e.g., primers labelled with biotin; (2) immobilizing the labelled PCR products on the walls of one or more capillary tubes having volumes on the order of 10-200 nanoliters; (3) preparing nanoliter quantities of labelled Sanger extension products of the amplified DNA; (4) purifying the oligonucleotide sequencing ladders; (5) high speed electrophoretic separation of the sequencing ladders; and (6) near-infrared, laser-induced fluorescence detection of the oligonucleotides. Base-calling is preferably performed in a single lane format with a single fluorophore, in which the bases are distinguished by different fluorescence lifetimes of dyes that otherwise have similar absorption and fluorescence emission spectra at the wavelengths used. Typical read lengths are on the order of 400-500 bases. Fluorescence is performed on-chip with one single-mode optical fiber carrying the excitation light to the capillary channel, and a second single-mode optical fiber collecting the fluorescent photons. Only sub-microliter volumes of expensive sequencing reagents and dye-labeled ddNTPs are required in this system.
REFERENCES:
patent: 5230781 (1993-07-01), Middendorf et al.
patent: 5360523 (1994-11-01), Middendorf et al.
patent: 5366603 (1994-11-01), Middendorf et al.
patent: 5374527 (1994-12-01), Grossman
patent: 5405746 (1995-04-01), Uhlen
patent: 5484701 (1996-01-01), Cocuzza et al.
Y. Xia, Soft Lithography: Micro- and Nanofabrication Based on Microcontact Printing and Teplica Molding, Engineeriing Materials Science, Diss. Abs. vol. 57, p. 6507-B (1997).
L. Amankwa et al., "Trypsin-Modified Fused-Silica Capillary Microreactor for Peptide Mapping by Capillary Zone Electrophoresis," Anal. Chem., vol. 64, pp. 1610-1613 (1992).
L. Amankwa et al., "On-Line Peptide Mapping by Capillary Zone Electrophoresis," Anal. Chem., vol. 65, pp. 2693-2697 (1993).
C. Effenhauser et al., "High-Speed Separation of Antisense Oligonucleotides on a Micromachined Capillary Electrophoresis Device," Anal. Chem., vol. 66, pp. 2949-2593 (1994).
Effenhauser et al., "Manipulation of Sample Fractions on a Capillary Electrophoresis Chip," Anal. Chem., vol. 67, pp. 2284-2287 (1995).
A. Wooley et al., "Ultra-High-Speed DNA Sequencing Using Capillary Electrophoresis Chips," Anal. Chem., vol. 67, pp. 3676-3680 (1995).
S. Jacobson et al., "Integrated Microdevice for DNA Restriction Fragment Analysis," Anal. Chem., vol. 68, pp. 720-723 (1996).
Z. Liang et al., "Microfabrication of a Panar Absorbance and Flourescence Cell for Integrated Capillary Electrophoresis Devices," Anal. Chem., vol. 68, pp. 1040-1046 (1996).
S. Hjerten "High-Performance Electrophoresis Elimination of Electroendosmosis and Solute Adsorption," J. Chromatogr., vol. 347, pp. 191-198 (1985).
A. Cohen et al., "Separation and Analysis of DNA Sequence Reaction Products by Capillary Gel Electrophoresis," J. Chromatogr., vol. 516, pp. 49-60 (1990).
S. Stahl et al., "Solid Phase DNA Sequencing Using the Biotin-Avidin System,"Nucl. Acid Res., vol. 16, pp. 3025-3037 (1988).
T. Hawkins et al., "DNA Purification and Isolation Using a Solid Phase," Nucl. Acid Res., vol. 22, pp. 4543-4544 (1994).
S. Huang et al., "Binding of Biotinylated DNA to Strepavidin-Coated Polystyrene Latex," Anal. Biochem., vol. 222, pp. 441-449 (1994).
A. Rolfs et al., "Fully-Automated, Nonradioactive Solid-Phase Sequencing of Genomic DNA Obtained from PCR," Biotechniques, vol. 17, pp. 782-787 (1994).
T. Hultman et al., "Direct Solid Phase Sequencing of Genomic and Plasmid DNA Using Magnetic Beads as Solid Supports," Nucl. Acid Res., vol. 17, pp. 4937-4945 (1989).
M. Uhlen et al., "Magnetic Separation of DNA," Nature, vol. 340, pp. 733-734 (1989).
D. Williams et al., "Single-Lane, Single-Fluor Sequencing using Dideoxy-Labled, Heavy-Atom Modified Near-IR Fluorescent Dyes," SPIE, vol. 2386, pp. 55-56 (proceedings paper, presentation in San Jose, California, Feb. 6, 1995).
D. Harrison et al., "Capillary Electrophoresis and Sample Injection Systems Integrated on a Planar Glass Chip," Anal.Chem., vol. 64, pp. 1926-1932 (1992).
Z. Fan et al., "Micromachining of Capillary Electrophoresis Injectors and Separators on Glass Chips and Evaluation of Flow at Capillary Intersections," Anal. Chem., vol. 66, pp. 177-184 (1994).
S. Jacobson et al., "Effects of Injection Schemes and Column Geometry on the Performance of Microchip Electrophoresis Devices," Anal. Chem., vol. 66, pp. 1107-1113 (1994).
S. Soper et al., "Micro-DNA Sequence Analysis Using Capillary Electrophoresis and Near-IR Fluorescence Detection," SPIE, vol. 2680, pp. 235-246 (proceedings paper, San Jose, California, Feb. 2, 1996).
D. Williams, slides presented at two meetings (Orlando, Florida, Jan. 24, 1996; and Baton Rouge, Louisiana, Mar. 16, 1996).
S. Soper et al., "On-Line Fluorescence Lifetime Determinations in Capillary Electrophoresis," poster (Orlando, Florida, Jan. 24, 1996).
S. Soper et al., "On-Line Fluorescence Lifetime Determinations in Capillary Electrophoresis," Anal. Chem., vol. 67, pp. 4358-4365 (Dec. 1, 1995).
D. Williams et al., "Ultrasensitive Near-IR Fluorescence Detection for Capillary Gel Electrophoresis and DNA Sequencing Applications," Anal. Chem., vol. 67, pp. 3427-3432 (Oct. 1, 1995).
S. Soper et al., "Multiplexed DNA Sequencing Using CGE and NIR Fluorescence," LSU Grant Application No. 5688, submitted to the Department of Health and Human Services, Public Health Service May 24, 1994.
S. Soper et al., "High Throughput DNA Sequencing Using Micro-Reactors and Micro-CE,"LSU Grant Application, submitted to the Department of Health and Human Services, Public Health Service, Aug. 22, 1995.
Davies Jack D.
Soper Steven A.
Vladimirsky Yuli
Arthur Lisa B.
Board of Supervisors of Louisiana State University and Agricultu
Runels John H.
Souaya Jehhanne
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