Imaging microarrays
First Claim
Patent Images
1. A method of detecting single copy number variation in a target spot of a genomic microarray comprising:
- acquiring an image of said microarray including said target spot;
processing the image to correct for chip misalignment;
processing the image to correct for background noise;
analyzing the image to identify a target patch;
analyzing the image to edit debris;
analyzing the image to correct for ratio bias;
detecting single copy number variation in the target spot by an objective statistical analysis;
wherein the genomic microarray includes a test genomic material marked with receptors having a first wavelength and a reference genomic material marked with receptors having a second wavelength, the test and reference genomic materials forming the target spot by a hybridization process;
further comprising measuring a fluorescent signal intensity of the target spot from the test genomic material and the reference genomic material, wherein the fluorescent signal intensity of the first wavelength is proportional to a copy number of the test genomic material and the fluorescent signal intensity of the second wavelength is proportional to a copy number of the reference genomic material;
obtaining an image of a theoretical set of patches; and
,cross-correlating the image of the theoretical set of patches to the image of the microarray.
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Abstract
A method of obtaining a corrected image of a microarray includes acquiring an image of a microarray including a target spot, and processing the image to correct for background noise and chip misalignment. The method also includes analyzing the image to identify a target patch, edit debris, and correct for ratio bias; and detecting single copy number variation in the target spot using an objective statistical analysis that includes a t-value statistical analysis. The method provides statistically robust computational processes for accurately detecting genomic variation at the single copy level.
24 Citations
30 Claims
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1. A method of detecting single copy number variation in a target spot of a genomic microarray comprising:
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acquiring an image of said microarray including said target spot; processing the image to correct for chip misalignment; processing the image to correct for background noise; analyzing the image to identify a target patch; analyzing the image to edit debris; analyzing the image to correct for ratio bias; detecting single copy number variation in the target spot by an objective statistical analysis; wherein the genomic microarray includes a test genomic material marked with receptors having a first wavelength and a reference genomic material marked with receptors having a second wavelength, the test and reference genomic materials forming the target spot by a hybridization process; further comprising measuring a fluorescent signal intensity of the target spot from the test genomic material and the reference genomic material, wherein the fluorescent signal intensity of the first wavelength is proportional to a copy number of the test genomic material and the fluorescent signal intensity of the second wavelength is proportional to a copy number of the reference genomic material; obtaining an image of a theoretical set of patches; and
,cross-correlating the image of the theoretical set of patches to the image of the microarray. - View Dependent Claims (2, 3, 4, 5, 6, 7, 13, 14)
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8. A method of detecting single copy number variation in a target spot of a genormic microarray comprising:
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acquiring an image of said microarray including said target spot; processing the image to correct for chip misalignment; processing the image to correct for background noise; analyzing the image to identify a target patch; analyzing the image to edit debris; analyzing the image to correct for ratio bias; detecting single copy number variation in the target spot by an objective statistical analysis; wherein the genomic microarray includes a test genomic material marked with receptors having a first wavelength and a reference genomic material marked with receptors having a second wavelength, the test and reference genomic materials forming the target spot by a hybridization process; further comprising measuring a fluorescent signal intensity of the target spot from the test genomic material and the reference genomic material, wherein the fluorescent signal intensity of the first wavelength is proportional to a copy number of the test genomic material and the fluorescent signal intensity of the second wavelength is proportional to a copy number of the reference genomic material; wherein correcting of the background noise includes; computing the acquired image; computing a minimum image based on the computed acquired image; computing a maximum image based on the computed minimum image; subtracting a background image from the computed acquired image to obtain a resulting image; and
,optimizing the correction for the background noise by identifying a background peak of the resulting image and subtracting the resulting image to obtain a mean value of zero for a corrected image. - View Dependent Claims (9, 10, 11, 12)
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15. A method of detecting single copy number variation in a target spot of a genomic microarray comprising:
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acquiring an image of said microarray including said target spot; processing the image to correct for chip misalignment; processing the image to correct for background noise; analyzing the image to identify a target patch; analyzing the image to edit debris; analyzing the image to correct for ratio bias; detecting single copy number variation in the target spot by an objective statistical analysis; wherein the genomic microarray includes a test genomic material marked with receptors having a first wavelength and a reference genomic material marked with receptors having a second wavelength, the test and reference genomic materials forming the target spot by a hybridization process; further comprising a fluorescent signal intensity of the target spot from the test genomic material and the reference genomic material, wherein the fluorescent signal intensity of the first wavelength is proportional to a copy number of the test genomic material and the fluorescent signal intensity of the second wavelength is proportional to a copy number of the reference genomic material;
further comprising;computing a ratio based on a test and a reference fluorescence signal intensity value associated with the target spot; further wherein correcting for ratio bias includes measuring a raw ratio representing a ratio of total test fluorescence intensity from the target spot and a total reference fluorescence intensity from the target spot; normalizing the raw ratio by dividing the raw ratio of each spot by the median over all spots of the raw ratio; and further wherein correcting for ratio bias includes; selecting a subset of spots that have a normalized observed ratio approximately equal to 1.0, referred to as “
plausibly modal spots”
;modeling an effect of bias in said subset of plausibly modal spots by the equation R=r−
B, where R represents said normalized observed ratio, r is the reference intensity of the spot, and B is a bias parameter that is constant across all spots of a microarray;estimating the optimum value of B from said subset of plausibly modal spots; performing ratio bias correction of unknown data by applying the correction formula Rcorrected=Roriginal/Rpredicted and choosing a value of B that minimizes the mean square difference between the set of normalized original ratios, and the set of normalized predicted ratios according to equation R=r−
B.
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16. A method of detecting single copy number variation in a target spot of a genomic microarray comprising:
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acquiring an image of said microarray including said target spot; processing the image to correct for chip misalignment; processing the image to correct for background noise; analyzing to identify a target patch; analyzing the image to edit debris; analyzing the image to correct for ratio bias; detecting single copy number variation in the target spot by an objective statistical analysis; wherein the genomic microarray includes a test genomic material marked with receptors having a first wavelength and a reference receptors having a second wavelength, the test and reference genomic materials forming the target spot by a hybridization process; further comprising measuring a fluorescent signal intensity of the target spot from the test genomic material and the reference genomic material, wherein the fluorescent signal intensity of the first wavelength is proportional to a copy number of the test genomic material and the fluorescent signal intensity of the second wavelength is proportional to a copy number of the reference genomic material; wherein the objective statistical analysis includes a target-modal statistical analysis; and wherein the target-modal statistical analysis includes; for a selected subset of plausibly modal targets, estimating a statistical distribution of ratios of plausibly modal targets; and for each target of the microarray, computing an estimated target mean and a lower 95% confidence interval limit; and for each target, reducing the lower 95% confidence interval limit by the median 95% confidence interval over all targets of the microarray; and for each target, calculating a confidence estimate. - View Dependent Claims (17)
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18. A computer program product residing on a computer storage medium having instructions stored thereon which, when executed by a logic processor, cause the processor to:
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acquire an image of a microarray including a target spot; process the image to correct for background noise and chip misalignment; analyze the image to detect target spots; analyze the image to identify the target patch, edit debris, and correct for ratio bias; detect single copy number variation in the target spot by an objective statistical analysis; wherein genomic microarray includes a test genomic material marked with receptors having a first wavelength and a reference genomic material marked with receptors having a second wavelength, the test and reference genomic materials forming the target spot by a hybridization process; measure a fluorescent signal intensity of the target spot from the test genomic material and the reference genomic material, wherein the fluorescent signal intensity of the first wavelength is proportional to a copy number of the test genomic material and the fluorescent signal intensity of the second wavelength is proportional to a copy number of the reference genomic material; obtain an image of a theoretical set of patches; and cross-correlate the image of the theoretical set of patches to the image of the microarray. - View Dependent Claims (19, 20, 21, 22, 23, 24, 25, 26, 27, 28)
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29. A processor and a memory configured to:
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acquire an image of a microarray including at least one target spot; process the image to correct for background noise and chip misalignment; analyze the image to detect target spots; analyze the image to identify a target patch, edit debris, and correct for ratio bias; and
,detect single copy number variation in a target spot by an objective statistical analysis; wherein the microarray includes a test genomic material marked with receptors having a first wavelength and a reference genomic material marked with receptors having a second wavelength, the test and reference genomic materials forming the target spot by a hybridization process; measure a fluorescent signal intensity of the target spot from the test genomic material and the reference genomic material, wherein the fluorescent signal intensity of the first wavelength is proportional to a copy number of the test genomic material and the fluorescent signal intensity of the second wavelength is proportional to a copy number of the reference genomic material; obtain an image of a theoretical set of patches; and
,cross-correlate the image of the theoretical set of patches to the image of the microarray.
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30. A system comprising:
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means for acquiring an image of a microarray including at least one target spot; means for processing the image to correct for background noise and chip misalignment; means to analyze the image to detect target spots; means for analyzing the image to identify a target patch, edit debris, and correct for ratio bias; and
,means for detecting single copy number variation in the target spot by an objective statistical analysis; wherein the microarray includes a test genomic material marked with receptors having a first wavelength and a reference genomic material marked with receptors having a second wavelength, the test and reference genomic materials forming the target spot by a hybridization process; means for measuring a fluorescent signal intensity of the target spot from the test genomic material and the reference genomic material, wherein the fluorescent signal intensity of the first wavelength is proportional to a copy number of the test genomic material and the fluorescent signal intensity of the second wavelength is proportional to a copy number of the reference genomic material; means for obtaining an image of a theoretical set of patches; and
,means for cross-correlating the image of the theoretical set of patches to the image of the microarray.
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Specification