Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Reexamination Certificate
2002-01-23
2004-04-27
Horlick, Kenneth R. (Department: 1634)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
C435S320100, C435S252800, C435S174000, C435S183000, C382S129000, C382S133000, C382S153000, C382S173000, C382S286000, C382S291000, C702S019000, C702S022000, C536S022100
Reexamination Certificate
active
06727068
ABSTRACT:
BACKGROUND
The field of genomics has taken rapid strides in recent years. It started with efforts to determine the entire nucleotide sequence of simpler organisms such as viruses and bacteria. As a result, genomic sequences of
Hemophilus influenzae
(Fleischman et al.,
Science
269: 496-512, 1995) and a number of other bacterial strains (
Escherichia coli, Mycobacterium tuberculosis, Helicobacter pylori, Caulobacter jejuni, Mycobacterium leprae
) are now available (reviewed in Nierman et al.,
Curr. Opin. Struct. Biol.
10: 343-348, 2000). This was followed by the determination of complete nucleotide sequence of a number of eukaryotic organisms including budding-yeast (
Saccharomyces cerevisiae
) (Goffeau et al.,
Science
274: 563-567, 1996), nematode (
Cenorhabditis elegans
) (
C. elegans
sequencing consortium,
Science
282: 2012-2018 1998) and fruit fly (
Drosophila melanogaster
) (Adams et al.,
Science
287: 2185-2195, 2000). The sequence of the human genome was published in February of 2001 (International Human Genome Sequence Consortium,
Nature,
409:860-921, 2001; Venter et al.,
Science,
291:1304-1351, 2001). Additionally, some of the ongoing efforts are currently focused on genome sequencing of agriculturally important plants such as rice (
Science
288: 239-240, 2000; Sasaki and Burr,
Curr. Opin. Plant Biol.
3: 138-141, 2000) and experimentally critical animal model such as mice (News Focus,
Science
288: 248-257, 2000).
The availability of complete genomic sequences of various organisms promises to significantly advance our understanding of various fundamental aspects of biology. It also promises to provide unparalleled applied benefits such as understanding genetic basis of certain diseases, providing new targets for therapeutic intervention, developing a new generation of diagnostic tests, etc. New and improved tools, however, will be needed to harvest and fully realize the potential of genomics research.
Even though the DNA complement or gene complement is identical in various cells in the body of multi-cellular organisms, there are qualitative and quantitative differences in gene expression in various cells. A human genome is estimated to contain roughly about 30,000-40,000 genes, however, only a fraction of these genes are expressed in a given cell (International Human Genome Sequence Consortium,
Nature,
409:860-921, 2001; Venter et al.,
Science,
291:1304-1351, 2001). Moreover, there are quantitative differences among the expressed genes in various cell types. Although all cells express certain housekeeping genes, each distinct cell type additionally expresses a unique set of genes. Phenotypic differences between cell types are largely determined by the complement of proteins that are uniquely expressed. It is the expression of this unique set of genes and their encoded proteins, which constitutes functional identity of a cell type, and distinguishes it from other cell types. Moreover, the complement of genes that are expressed, and their level of expression vary considerably depending on the developmental stage of a given cell type. Certain genes are specifically activated or repressed during differentiation of a cell. The level of expression also changes during development and differentiation. Qualitative and quantitative changes in gene expression also take place during cell division, e.g. in various phases of cell cycle. Signal transduction by biologically active molecules such as hormones, growth factors and cytokines often involves modulation of gene expression. Global change in gene expression also plays a determinative role in the process of aging.
In addition to the endogenous or internal factors mentioned above, certain external factors or stimuli, such as environmental factors, also bring about changes in gene expression profiles. Infectious organisms such as bacteria, viruses, fungi and parasites interact with the cells and influence the qualitative and quantitative aspects of gene expression. Thus, the precise complement of genes expressed by a given cell type is influenced by a number of endogenous and exogenous factors. The outcome of these changes is critical for normal cell survival, growth, development and response to the environment. Therefore, it is important to identify, characterize and measure changes in gene expression. The knowledge gained from such analysis will not only further our understanding of basic biology, but it will also allow us to exploit it for various purposes such as diagnosis of infectious and non-infectious diseases, screening to identify and develop new drugs, etc.
Besides the conventional, one by one gene expression analysis methods like Northern analysis, RNase protection assays, and real time PCT (RT-PCR); there are several methods currently available to examine gene expression in a genome wide scale. These approaches are variously referred to as RNA profiling, differential display, etc. These methods can be broadly divided into three categories: (1) hybridization-based methods such as subtractive hybridization (Koyama et al.,
Proc. Natl. Acad. Sci. USA
84: 1609-1613, 1987; Zipfel et al.,
Mol. Cell. Biol.
9: 1041-1048, 1989), microarray, etc., (2) cDNA tags: EST, serial analysis of gene expression (SAGE) (see, e.g. U.S. Pat. Nos. 5,695,937 and 5,866,330), and (3) fragment size based, often referred to as gel-based methods where a differential display is generated upon electrophoretic separation of DNA fragments on a gel such as a polyacrylamide gel (described in U.S. Pat. Nos. 5,871,697, 5,459,037, 5,712,126 and PCT publication No. WO 98/51789).
Expressed Sequence Tags (ESTs) are created by partially sequencing (usually a single pass) randomly chosen gene transcripts that have been converted into cDNA. The concept of ESTs as an alternative to genome sequencing for the rapid determination of the expressed complement of a genome was first introduced by Adams et al. (
Science
252: 1651-1656, 1991). The EST approach, combined with the power of high throughput sequencing, has brought about a revolution (Zweiger and Scott,
Curr. Opin. Biotechnol.
8: 684-687, 1997). EST tags often contain enough information to identify the transcript corresponding to the cDNA clone by searching nucleotide and protein databases. Thus, the EST approach provides a valuable tool for the identification and characterization of novel genes. ESTs are also long enough to be reliably used as substrate for microarrays, and are less expensive and superior in performance than oligonucleotides for the purpose. The use of EST-based microarray technology is finding increasing use in global genome wide transcript profiling. The number of EST sequences being deposited in databanks is exponentially growing. A sub-database containing EST sequences (dbEST) is available in GenBank. A large number of EST sequences from various organisms are also available in public databanks.
The National Cancer Institute (NCI) launched the first large-scale EST project in 1997, the Cancer Genome Anatomy Program (CGAP), focusing on a single aspect, the comprehensive molecular characterization of human normal, pre-cancerous and malignant cells (Strausberg et al.,
Trends Genet.
16: 103-106, 2000). Similarly, tissue-specific EST databases have also been created in order to understand the unique physiological functions of a tissue/organ as well as to gain insight into changes into global gene expression associated with various pathological conditions. For instance, a collection of EST sequences derived from human heart tissue of various anatomical, developmental and pathological stages has been established with a view to understand cardiovascular diseases (Dempsey et al.,
Mol. Med. Today,
6: 231-237, 2000). The EST profile of an organ typically reflects the unique function of the organ. For example, the pancreas and liver have a large proportion of ESTs corresponding to secreted proteins, whereas the brain and heart have a very small proportion of such ESTs. Similarly, ESTs corresponding to contractile proteins are present in high proportion in the heart,
Li Bi-Yu
Shi Liang
Wang Xun
Horlick Kenneth R.
Kakefuda Mary
Syngenta Participations (AG)
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