Chemical apparatus and process disinfecting – deodorizing – preser – Analyzer – structured indicator – or manipulative laboratory... – Means for analyzing gas sample
Reexamination Certificate
2000-02-25
2003-07-01
Warden, Jill (Department: 1743)
Chemical apparatus and process disinfecting, deodorizing, preser
Analyzer, structured indicator, or manipulative laboratory...
Means for analyzing gas sample
C422S068100, C422S082050, C422S082060, C422S105000, C422S105000, C435S283100, C435S287100, C435S287800, C435S288200, C435S288300, C435S288400, C436S174000, C436S180000, C264S239000, C264S241000, C264S250000, C264S259000, C264S297100, C264S297400
Reexamination Certificate
active
06585939
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to methods and compositions for performing biological assays. Specifically, the invention provides compositions, referred to herein as microstructures, which may be used in micro-fabricated devices for a variety of biological applications, including nucleic acid sequence analysis and polynucleotide amplification, as well as other biological assays and reactions.
BACKGROUND OF THE INVENTION
Many techniques and assays have been developed for the analysis of biological samples. Practical applications of these techniques include the diagnosis of genetic diseases, the diagnosis of infectious diseases, forensic techniques, paternity determination, and genome mapping.
For example, in the field of nucleotide sequence analysis, many techniques have been developed to analyze nucleic acid sequences and detect the presence or absence of various genetic elements such as genetic mutations, and polymorphisms such as single nucleotide polymorphisms(hereinafter “SNPs”), base deletions, base insertions, and heterozygous as well as homozygous polymorphisms.
Currently, the most definitive method for analyzing nucleic acid sequences is to determine the complete nucleotide sequence of each nucleic acid segment of interest. Examples of how sequencing has been used to study mutations in human genes are included in the publications of Engelke et al. (1998
, Proc. Natl. Acad. Sci. U.S.A.
85:544-548) and Wong et al. (1997
, Nature
300:384-386). The most commonly used methods of nucleic acid sequencing include the dideoxy-mediated chain termination method, also known as the “Sanger Method” (Sanger, F. et al., 1975,
J. Molec. Biol.
94:441; Porbe, J. et al., 1997, Science 238:336-340) and the chemical degradation or “Maxam-Gilbert” method (Maxam, A. M. et al., 1977, Proc. Natl. Acad. Sci. U.S.A. 74:560).
Restriction fragment length polymorphism (hereinafter “RFLP”) mapping is another commonly used screen for DNA polymorphisms arising from DNA sequence variation. RFLP consists of digesting DNA with restriction endonucleases and analyzing the resulting fragments, as described by Botstein et al. (1980, Am. J. Hum. Genet. 32:314-331) and by White et al. (1998, Sci. Am. 258:40-48). Mutations that affect the sequence recognition of the endonuclease will alter enzymatic cleavage at that site, thereby altering the cleavage pattern of the DNA. DNA sequences are compared by looking for differences in restriction fragment lengths.
However, sequencing techniques such as the Sanger and Maxam-Gilbert methods involve series of nested reactions which are then analyzed on electrophoretic gels. RFLP analysis also requires analysis of reaction products on Southern Blots. These techniques can therefore be cumbersome to perform and analyze.
Alternative, simpler, and less cumbersome methods to analyze and/or sequence nucleic acid molecules have also been proposed. For example, there is considerable interest in developing methods of de novo sequencing using solid phase arrays (see, e.g., Chetverin, A. B. et al., 1994, Biotech. 12:1093-1099; Macevicz, U.S. Pat. No. 5,002,867; Beattie, W. G. et al., 1995, Molec. Biotech. 4:213-225; Drmanac, R. T., EP 797683; Church et al., U.S. Pat. No. 5,149,625; Gruber, L. S., EP 787183; each of which is incorporated herein by reference in its entirety) including universal sequencing arrays such as those described, e.g., by Head, S. et al. (U.S. patent application Ser. No. 08/976,427, filed Nov. 21, 1997, which is incorporated herein by reference in its entirety) and by Boyce-Jacino, M. et al. (U.S. patent application Ser. No. 09/097,791, filed Jun. 16, 1998, which is incorporated herein by reference in its entirety).
Other methods have been developed which use solid phase arrays to analyze single nucleotide polymorphisms (SNPs). For example, several primer-guided nucleotide incorporation procedures for assaying polymorphic sites in DNA have been described (e.g., Kornher, J. S. et al., 1989, Nucl. Acids Res. 17:7779-7784; Sokolov, B. P., 1990, Nucl. Acids Res. 18:3671; Syvanen, A.-C. Et al., 1990, Genomics 8:684-692; Kuppuswamy, M. N. et al., 1991, Proc. Natl. Acad. Sci. U.S.A. 88:11431147; Prezant, T. R. et al., 1992, Hum. Mutat. 1:159-164; Ugozzoli, L. et al., 1992, GATA 9:107-112; Nyren, P. et al., 1993, Anal.
Biochem.
208:171-175; and Wallace WO89/10414).
An alternative “micro sequencing” method, the Genetic Bit Analysis (GBA′″) method has been disclosed by Goelet, P. et al. (WO 92/15712). Several other micro sequencing methods have also been described, including variations of the (GBA
TM
) 10 method of Goelet et al. (see, e.g., Mundy, U.S. Pat. No. 4,656, 127; Vary and Diamond, U.S. Pat. No. 4,851, 331; Cohen, D. et al., PCT Application No. WO 91/02087; Chee, M. et al., PCT Application No. WO 95/11995; Landegren, U. et al., 1998, Science 241:1077-1080; Nicerson, D. A. et al., 1990, Proc. Natl. Acad. Sci. U.S.A. 87:8923-8927; Pastinen, T. et al., 1997, Genome Res. 7:606-614; Pastinen, T. et al., 1996, Clin. Chem. 42:1391-1397; Jalanko, A. et al., 1992, Clin. Chem. 38:39-43; Shumaker, J. M. et al., PCT Application Wo 95/00669).
Other methods of nucleic acid analysis involve amplifying defined segments of nucleic acid sequence for subsequent analysis, e.g., by one or more of the micro sequencing methods discussed above. For example, the Polymerase Chain Reaction (PCR) is widely used to amplify defined segments of nucleic acid sequence in vitro. Generally, a targeted polynucleotide segment is flanked by two oligonucleotide primers. PCR consists of three steps that are repeated many times in a cyclical manner: (1) denaturing double-stranded polynucleotide sample at high temperature (about 94° C.); (2) annealing oligonucleotide primers to the polynucleotide template at low temperature (from about 37° C. to about 55° C.); and (3) extending primers using a template-dependent polymerase at a moderate temperature (about 72° C.).
PCR has been demonstrated in micro-fabricated devices consisting of a reaction chamber that has been etched into a silicon chip (see, e.g., Wilding et al., 1994,
Clin. Chem.
40:1815-1818; Northrup et al., 1993, Transducers' 93:924-926), and continuous flow PCR has been accomplished on a chip (see, e.g., Kopp et al., 1998, Science 280:1046-1048). However, these devices have the major drawback of significant solution evaporation at the high temperatures used in PCR. Further, the devices do not contain integrated micro valves or micro pumps.
A powerful concept and method known as Microfluidicsenabled Target Amplification (MeTA), which may be used as an alternative to PCR, has been previously described as set forth in U.S. patent application Ser No. 08/924,763 (Kumar, R., “AMPLIFICATION METHOD FOR A POLYNUCLEOTIDE,” filed Aug. 27, 1997) which is incorporated herein, by reference, in its entirety. This target amplification method is an isothermal process, using chemicals rather than high temperature DNA denaturation. MeTA method is preferably implemented using a micro-fabricated device such as the device disclosed in U.S. provisional application No. 60/110,367 (Fan, Z. H. et al., “MICROFLUIDICS-BASED DEVICE FOR DNA TARGET AMPLIFICATION” filed Nov. 30, 1998) which is incorporated herein, by reference, in its entirety.
However, implementation of such methods requires special micro-fabricated devices. Currently, such micro-fabricated devices are typically fabricated from glass, silicon, or plastic plates or slides, which may be etched with horizontal or vertical cells (chambers) and/or channels, e.g., by photolithography, chemical and/or laser etching, and bonding techniques. However, such devices are fragile and expensive to produce. Alternatively, techniques have been developed for fabricating surface relief patterns in the plane of self-assembled monolayers (“SAMs”). These techniques typically comprise casting a material such as polydimethylsiloxane (PDMS), in the form of a prepolymer, onto a complementary relief pattern (i.e., a “cast”) and then curing the prepolymer (see, e.g., Wilbur et al., 1995,
A
Handy Dwayne K.
Mierzwa Kevin G.
Orchid BioSciences, Inc.
Warden Jill
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