C - Chemistry – Metallurgy – 12 – Q
Patent
C - Chemistry, Metallurgy
12
Q
C12Q 1/68 (2006.01) G01N 33/48 (2006.01) G01N 33/50 (2006.01) G06F 19/00 (2006.01)
Patent
CA 2327527
High-throughput measurement of hybridization signatures obtained using complex probes prepared from total RNA, DNA microarray and two-color fluorescence detection requires accurate normalization based on reliable internal control. The invention relates to rRNA cDNA probes used as internal control to achieve normalization of fluorescence detection. Analysis of data obtain from laser scanner during DNA microarray experiment first needs image processing. The data generated for the arrayed genes must be further normalized before differentially expressed genes can be identified. Normalization is necessary to adjust for differences in labeling and detection efficiencies for the fluorescent labels and for differences in the quantity of starting RNA from the two samples examined in the assay. These problems can cause a shift in the average ratio of the two fluorophor (Cy3 and Cy5 as an example) and the intensities must be resealed before an experiment can be properly analyzed. Because of its invariant expression across tissues and treatments, 18S and 28S RNA are ideal internal control for quantitative RNA analysis. However the high abundance of these rRNA makes it difficult to use in microarray experiments with mRNA species that are less abundant. We solve this problem by the utilization of specific amount of competitive complementary oligonucleotide during the hybridization step to adjust the signal detection of these rRNA cDNA to less abundant cDNA. The invention may be used for screening of differentially expressed gene expression with cDNA microarray or oligo microarray (DNA microarray).
Camato Rino N.
Larose Anne-Marie
Geneka Biotechnologie Inc.
Goudreau Gage Dubuc
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