Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Preparing compound containing saccharide radical
Reexamination Certificate
2000-01-05
2003-03-18
Whisenant, Ethan C. (Department: 1655)
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Preparing compound containing saccharide radical
C435S006120, C435S091100, C435S287100, C435S287200, C436S094000, C536S023100
Reexamination Certificate
active
06534293
ABSTRACT:
FIELD OF THE INVENTION
The present invention is directed to accelerating identification of single nucleotide polymorphisms and an alignment of clone in genomic sequencing.
BACKGROUND OF THE INVENTION
Introduction to Applications of SNPS
Accumulation of genetic changes affecting cell cycle control, cell differentiation, apoptosis, and DNA replication and repair lead to carcinogenesis (Bishop, J. M., “Molecular Themes In Oncogenesis,”
Cell
, 64(2):235-48 (1991)). DNA alterations include large deletions which inactivate tumor supressor genes, amplification to increase expression of oncogenes, and most commonly single nucleotide mutations or polymorphisms which impair gene expression or gene function or predispose an individual to further genomic instability (Table 1).
TABLE 1
Genetic Alterations Commonly Found in thc Human Genome
Possible
Type of
Possible Causes
Consequences
Detection
Alteration
of Alteration
of Alteration
of Alteration
Single
Inherited variation
Silent: does not
DNA
nucleotide
Methylation
alter function
sequencing
polymorphism
Carcinogens
Missense: alters
SSCP,
(SNP)
Defective repair
gene function
DGGE,
genes
Nonsense: truncates
CDGE
gene
Protein
truncation
Mismatch
cleavage
Microsatellite
Defective DNA
Frameshift:
Microsatel-
instability
repair genes
truncates gene
lite Analysis
(MIN)
Carcinogens
Large deletions
Defective DNA
Loss of gene
Loss of
repair genes
function
hetero-
Defective DNA
zygosity
replication genes
CGH
Illegitimate
SNP analysis
recombination
Double strand break
DNA
Defective DNA
Overexpression
Competitive
amplifications
repair genes
of gene
PCR
Defective DNA
CGH
replication genes
SNP analysis
Illegitimate
recombination
Others:
Defective methylase
Gene silencing
Endonu-
Methylation,
genes
or overexpression;
clease
Translocation
Double strand break
creation of
digestion
chimeric protein
PCR, FISH
Rapid detection of germline mutations in individuals at risk and accurate characterization of genetic changes in individual tumors would provide opportunities to improve early detection, prevention, prognosis, and specific treatment. However, genetic detection poses the problem of identifying a predisposing polymorphism in the germline or an index mutation in a pre-malignant lesion or early cancer that may be present at many potential sites in many genes. Furthermore, quantification of allele copy number is necessary to detect gene amplification and deletion. Therefore, technologies are urgently needed that can rapidly detect mutation, allele deletion, and allele amplification in multiple genes. Single nucleotide polymorphisms (“SNP”s) are potentially powerful genetic markers for early detection, diagnosis, and staging of human cancers.
Identification of DNA sequence polymorphisms is the cornerstone of modern genome mapping. Initially, maps were created using RFLP markers (Botstein, D., et al., “Construction Of A Genetic Linkage Map In Man Using Restriction Fragment Length Polymorphisms,”
Amer. J. Hum. Genet
., 32:314-331 (1980)), and later by the more polymorphic dinucleotide repeat sequences (Weber, J. L. et al., “Abundant Class Of Human DNA Polymorphisms Which Can Be Typed Using The Polymerase Chain Reaction,.”
Amer. J. Hum. Genet
., 44:388-396 (1989) and Reed, P. W., et al., “Chromosome-Specific Microsatellite Sets For Fluorescence-Based, Semi-Automated Genome Mapping,”
Nat Genet
, 7(3): 390-5 (1994)). Such sequence polymorphisms may also be used to detect inactivation of tumor suppressor genes via LOH and activation of oncogenes via amplification. These genomic changes are currently being analyzed using conventional Southern hybridizations, competitive PCR, real-time PCR, microsatellite marker analysis, and comparative genome Hybridization (CGH) (Ried, T., et al., “Comparative Genomic Hybridization Reveals A Specific Pattern Of Chromosomal Gains And Losses During The Genesis Of Colorectal Tumors,”
Genes, Chromosomes
&
Cancer
, 15(4):234-45 (1996), Kallioniemi, et al., “ERBB2 Amplification In Breast Cancer Analyzed By Fluorescence In Situ Hybridization,”
Proc Natl Acad Sci USA
, 89(12):5321-5 (1992), Kallioniemi, et al., “Comparative Genomic Hybridization: A Rapid New Method For Detecting And Mapping DNA Amplification In Tumors,”
Semin Cancer Biol
, 4(1):41-6 (1993), Kallioniemi, et al., “Detection And Mapping Of Amplified DNA Sequences In Breast Cancer By Comparative Genomic Hybridization,”
Proc Natl Acad Sci USA
, 91(6):2156-60 (1994), Kallioniemi, et al., “Identification Of Gains And Losses Of DNA Sequences In Primary Bladder Cancer By Comparative Genomic Hybridization,”
Genes Chromosom Cancer
, 12(3):213-9 (1995), Schwab, M., et al., “Amplified DNA With Limited Homology To Myc Cellular Oncogene Is Shared By Human Neuroblastoma Cell Lines And A Neuroblastoma Tumour,”
Nature
, 305(5931):245-8 (1983), Solomon, E., et al., “Chromosome 5 Allele Loss In Human Colorectal Carcinomas,”
Nature
, 328(6131):616-9 (1987), Law, D. J., et al., “Concerted Nonsyntenic Allelic Loss In Human Colorectal Carcinoma,”
Science
, 241(4868):961-5 (1988)., Frye, R. A., et al., “Detection Of Amplified Oncogenes By Differential Polymerase Chain Reaction,” Oncogene, 4(9):1153-7 (1989), Neubauer, A., et al., “Analysis Of Gene Amplification In Archival Tissue By Differential Polymerase Chain Reaction,”
Oncogene
, 7(5):1019-25 (1992), Chiang, P. W., et al., “Use Of A Fluorescent-PCR Reaction To Detect Genomic Sequence Copy Number And Transcriptional Abundance,”
Genome Research
, 6(10):1013-26 (1996), Heid, C. A., et al., “Real Time Quantitative PCR,”
Genome Research
, 6(10):986-94 (1996), Lee, H. H., et al., “Rapid Detection Of Trisomy 21 By Homologous Gene Quantitative PCR (HGQ-PCR),”
Human Genetics
, 99(3):364-7 (1997), Boland, C. R., et al., “Microallelotyping Defines The Sequence And Tempo Of Allelic Losses At Tumour Suppressor Gene Loci During Colorectal Cancer Progression,”
Nature Medicine
, 1(9):902-9 (1995), Cawkwell, L., et al., “Frequency Of Allele Loss Of DCC, p53, RB1, WT1, NF1, NM23 And APC/MCC In Colorectal Cancer Assayed By Fluorescent Multiplex Polymerase Chain Reaction,”
Br J Cancer
, 70(5):813-8 (1994), and Hampton, G. M., et al., “Simultaneous Assessment Of Loss Of Heterozygosity At Multiple Microsatellite Loci Using Semi-Automated Fluorescence-Based Detection: Subregional Mapping Of Chromosome 4 In Cervical Carcinoma,”
Proceedings of the National Academy of Sciences of the United States of America
, 93(13):6704-9 (1996)). Competitive and real-time PCR are considerably faster and require less material than Southern hybridization, although neither technique is amenable to multiplexing. Current multiplex microsatellite marker approaches require careful attention to primer concentrations and amplification conditions. While PCR products may be pooled in sets, this requires an initial run on agarose gels to approximate the amount of DNA in each band (Reed, P. W., et al., “Chromosome-Specific Microsatellite Sets For Fluorescence-Based, Semi-Automated Genome Mapping,”
Nat Genet
, 7(3): 390-5 (1994), and Hampton, G. M., et al., “Simultaneous Assessment Of Loss Of Heterozygosity At Multiple Microsatellite Loci Using Semi-Automated Fluorescence-Based Detection: Subregional Mapping Of Chromosome 4 In Cervical Carcinoma,”
Proc. Nat'l. Acad. Sci. USA
, 93(13):6704-9 (1996)). CGH provides a global assessment of LOH and amplification, but with a resolution range of about 20 Mb. To improve gene mapping and discovery, new techniques are urgently needed to allow for simultaneous detection of multiple genetic alterations.
Amplified fragment length polymorphism (“AFLP”) technology is a powerful DNA fingerprinting technique originally developed to identify plant polymorphisms in genomic DNA. It is based on the selective amplification of restriction fragments from a total digest of genomic DNA.
The original technique involved three steps: (1) restriction of the genomic DNA, i.e. with EcoRI and MseI, and ligation of oligonucleotide adapters, (2) selective amplification of a subset of all the fragments in the total digest using primers which reached in b
Barany Francis
Gerry Norman P.
Kirk Brian W.
Liu Jianzhao
Paty Philip B.
Cornell Research Foundation Inc.
Lu Frank W
Nixon & Peabody LLP
Whisenant Ethan C.
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