Data processing: measuring – calibrating – or testing – Measurement system in a specific environment – Biological or biochemical
Reexamination Certificate
2001-06-18
2002-09-17
Brusca, John S. (Department: 1637)
Data processing: measuring, calibrating, or testing
Measurement system in a specific environment
Biological or biochemical
C435S006120, C435S091100, C536S023100
Reexamination Certificate
active
06453243
ABSTRACT:
PRIORITY INFORMATION
This application claims priority to Japanese Application Serial No. 265933/2000, filed Sep. 1, 2000.
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to display and evaluation of gene expression data that are obtained by hybridizing genes to a particular gene with known identity. The present invention also relates to a method for displaying and evaluating failures, or errors, occurring in experimental processes for obtaining such data in a manner that is visually easy to interpret.
2. Description of the Related Art
As the number of biological species increases whose genome have been sequenced, genome comparison analyses have become widely used to find genes that evidence evolution of species and search for gene populations that are common among different species. Gene comparison is also employed to find any clues from the differences between species to identify characteristics specific to a particular species.
Due to the recent developments of technological infrastructures such as biochips or DNA chips (which are referred to as “biochips,” hereinafter), the subject of interest in molecular biology have been shifting from interspecific information to intraspecific information, namely, simultaneous expression analyses. This type of information, together with conventional interspecific comparisons, widens the possibility of the art from merely extracting information to associating pieces of the information with each other.
For example, if an unknown gene is found to have an expression pattern identical to that of a known gene, it is inferred that the unknown gene has a similar function to the known gene. Functions of these genes and the resulting proteins are studied by considering them as a functional unit or group. Further, how genes or proteins interact with each other is analyzed by associating them with the data for a known enzyme reaction or metabolism, or more directly, by making a gene deficit to terminate the expression of the gene or by making the gene excessively active to permit the overexpression and studying direct or indirect influences of the gene on expression patterns of the entire genes.
In studies of gene expression patterns using biochips, elements that are associated with living tissue of interest are prepared. The term “elements” herein refers to fragments of any DNA that are related to the living tissue of interest. In a biochip, the elements are spotted and immobilized on a substrate such as a slide glass or a silicon wafer with a density of several hundred to several thousand elements per square centimeter. The term “sample” herein refers to fragments of any DNA or RNA that are extracted from living tissue of interest to be reacted with the elements on a biochip. When a gene is expressed in cells, DNA is transcribed into RNA. The RNA is extracted and labeled with a fluorescent marker to serve as a sample. When a sample is reacted with an element, single strands that are complementary to each other bind, or hybridize, to one another. Thus, biochips permit quantitative or qualitative analyses of gene expressions in living tissue by taking advantage of hybridization.
A successful example in the art is the experiment conducted by University of Tokyo, Institute of Medical Science with regard to drug efficacy (T. Tsunoda et al.: Discrimination of Drug Sensitivity of Cancer Using cDNA Microarray and Multivariate Statistical Analysis: Genome informatics 1999 (December 1999) pp.227-228, Universal Academy Press Inc.). In this experiment, RNA extracted from normal cells and RNA extracted from cancer cells are each labeled with a fluorescent dye of different colors. The two types of RNA were mixed and allowed to hybridize to elements (i.e., genes) on a biochip. The intensities of fluorescent signals emitted from each of the two fluorescent dyes were measured.
FIG. 16
schematically shows the manner in which the state of each gene expression that has been obtained from the above-described experiment is displayed. In this manner of display, the data for fluorescent signals resulting from hybridization with genes immobilized on a biochip are plotted on a graph, with one axis representing the fluorescent signals for normal cells and the other representing the signals for cancer cells. One point in the graph corresponds to one gene. In analyzing data, among genes that emit fluorescent signals with higher intensities than a predetermined value, those that are specific to disease conditions are discriminated against the other genes on the basis of the ratio of the signal intensity for the normal cells to the signal intensity for the cancer cells. Specifically, genes corresponding to the points in the area A (i.e., genes that function in normal cells but not in cancer cells) and genes corresponding to the points in the area B (i.e., genes that function in cancer cells but not in normal cells) in
FIG. 16
are particularly distinguished. In this manner of displaying data, genes that function specifically in a specific disease can be discriminated.
The data used in such data analysis must be sufficiently reliable in itself to ensure feasibility of the analysis. In other words, the results should be reproducible in experiments conducted under the same conditions. However, the actual manufacturing technologies of biochips, as well as the techniques required for conducting experiments using biochips, are yet to be fully developed, and the reproducibility of experiments is not fully ensured. Underlying causes for this include the difficulty in spotting exactly equal amounts of elements on a biochip and the susceptibility of the technology to changes in environmental factors such as temperatures and humidity. Furthermore, the techniques have not been fully established to ensure constant hybridization reaction rates and the accuracy of the readings of fluorescent light after hybridization. At present, there is a considerable uncertainty concerning the reliability of the data obtained from these experiments.
FIG. 17
schematically shows an image data obtained when the results of a biochip experiment are read by a scanner. Until now, researchers have needed to visually examine such read images of biochips to determine if the data are usable or not. For example, data for a biochip is determined to be unusable when the read image data is dark throughout it (i.e., no expression is observed.), or when the image is partially bright (i.e., incomplete expression). These conditions seem to occur such as when hybridization is incomplete or when the substrate of the biochip is scratched or when spotted amounts on the biochip are not uniform throughout the biochip, though the exact causes are not known.
At present, from manufacturers' point of view, there is an increasing need for technologies to improve the accuracy of manufacturing processes of biochips and to enable mass production of reliable biochips with decreased errors. Thus, proper evaluation methods or tools are needed to accurately determine the accuracy and errors in the manufacturing of biochips. In contrast, from the researchers' point of view who use the biochip in their experiments, it will be convenient if proper evaluation methods or tools are provided for evaluating the results of biochip experiments in order to allow the user to determine if the results are usable or not, and if not, allow the user to find out the exact cause of it. Thus, a need exists for evaluation methods that enable the user to know what faulty events have taken place at what point of the manufacturing process of biochips and/or experiments using biochips and take into account the results in the later manufacturing or experiments.
SUMMARY OF THE INVENTION
The present invention addresses such a need of both of biochip manufacturers and users. Accordingly, it is an object of the present invention to provide effective methods for detecting any faulty events in the manufacturing process of biochips or in experiments using biochips from the data obtained in the experiments using the biochips.
Nakashige Ryo
Nozaki Yasuyuki
Tamura Takuro
Watanabe Tsunehiko
Brusca John S.
Hitachi Software Engineering Co, Ltd.
Kim Young
Pillsbury & Winthrop LLP
Wetherell, Jr. John R.
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