Use of representations of DNA for genetic analysis

Abstract

It is an object of the present invention to provide a solution to problems associated with the use of microarray technology for the analysis DNA. The present invention provides compositions and methods for the use of simple and compound representations of DNA in microarray technology. The present invention is also directed to methods for the production of High Complexity Representations (HCRs) of the DNA from cells.

Description

[0001] This application claims the benefit of International Patent Application No. PCT / US98 / 23168, which claims benefit of U.S. Provisional Application No. 60 / 064,358, filed 30 Oct. 1997, each of which is incorporated herein in its entirety.1. FIELD OF THE INVENTION[0003] The field of the invention is genetic analysis.2. BACKGROUND OF THE INVENTION2.1. MICROARRAY TECHNOLOGY[0004] Although global methods for genomic analysis, such as karyotyping, determination of ploidy, and more recently comparative genomic hybridizaton (CGH) (Feder et al.,1998, Cancer Genet. Cytogenet. 102:25-31; Gebhart et al., 1998, Int. J. Oncol. 12:1151-1155; Larramendy et al., 1997, Am. J. Pathol. 151:1153-1161; Lu et al., 1997, Genes Chromosomes Cancer 20:275-281, all of which are incorporated herein by reference) have provided useful insights into the pathophysiology of cancer and other diseases or conditions with a genetic component, and in some instances have aided diagnosis, prognosis and selection of tre...

AI Technical Summary

Benefits of technology

0022] Representation of the genome results in a simplification of its complexity; the genomic complexity of a representation can range from below 1% to as high as 95% of the total genome. This simplification allows for desirable hybridization kinetics. Probes from representations of genomic DNA can be used as the probe of the microarray, and as the labeled sample hybridized to any microarray, however derived. Because formation of a representation involves the step of amplifying the DNA via an amplification reaction, such as the polymerase chain reaction, ligase chain reaction, etc., very

Problems solved by technology

62:676-689; Gebhart et al., 1998, Int. J. Oncol. 12:1151-1155; Hacia et al., 1996, Nat. Genet. 14:441-447, all of which are incorporated herein by reference), this one-by-one query is an inefficient and incomplete method for genetically typing cells.

This method is very likely to yield useful information about cancer, but suffers limitations.

First, the interpretation of the data obtained and its correlation with disease process is likely to be a complex and difficult problem: multiple changes in gene expression will be observed that are not relevant to the disease of interest.

Second, our present cDNA collections are not complete, and any chip is likely to be obsolete in the near future.

Third, while a picture of the current state of the cell might be obtained, there would be little direct information about how the cell arrived at that state.

Lastly, obtaining reliable mRNA from biopsies is likely to be a difficult problem, because RNA is very unstable and undergoes rapid degradation due to the presence of ubiquitous RNAses.

Because a simple probe array is generally able to capture only a single species of DNA from the sample, this detection threshold poses a problem for the use of simple DNA probe arrays for analysis of genomic DNA.

Hence, in its unaltered format, the simple DNA probe chip would not suffice for the robust detection of genomic sequences.

Because each address contains fragments derived from the entire BAC clone, several problems are created.

Also, the great size of the megacloning vector inserts limits the positional resolution.

Another drawback is the presence of DNA derived from the megacloning vector and host sequences.

The steps of excisi

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