Methods and computer software for detecting splice variants
First Claim
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1. A computer-implemented method for identifying exons that are differentially spliced between a first and second sample comprising:
- (a) obtaining an exon intensity measurement for a first exon in a first gene in said first sample and in said second sample;
(b) obtaining a gene level measurement for said first gene in said first sample and in said second sample;
(c) obtaining a first normalized intensity measurement for said first exon in said first sample and a second normalized intensity measurement for said first exon in said second sample by dividing the intensity measurements obtained in (a) by the gene level measurements obtained in step (b) wherein the exon intensity in said first sample is divided by the gene level measurement in said first sample and the exon intensity in said second sample is divided by the gene level measurement in said second sample;
(d) calculating a splicing index measurement for said first exon, wherein the splicing index is equal to the log2 of the normalized intensity for said first exon in said first sample divided by the normalized intensity for said first exon in said second sample; and
(e) identifying said first exon as being differentially spliced if the absolute value of the splicing index obtained in (d) is greater than a threshold value.
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Abstract
Methods and software products for analysis of alternative splicing are disclosed. In general the methods involve normalizing probe set or exon intensity to an expression level measurement of the gene. The methods may be used to identify tissue-specific alternative splicing events.
55 Citations
15 Claims
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1. A computer-implemented method for identifying exons that are differentially spliced between a first and second sample comprising:
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(a) obtaining an exon intensity measurement for a first exon in a first gene in said first sample and in said second sample;
(b) obtaining a gene level measurement for said first gene in said first sample and in said second sample;
(c) obtaining a first normalized intensity measurement for said first exon in said first sample and a second normalized intensity measurement for said first exon in said second sample by dividing the intensity measurements obtained in (a) by the gene level measurements obtained in step (b) wherein the exon intensity in said first sample is divided by the gene level measurement in said first sample and the exon intensity in said second sample is divided by the gene level measurement in said second sample;
(d) calculating a splicing index measurement for said first exon, wherein the splicing index is equal to the log2 of the normalized intensity for said first exon in said first sample divided by the normalized intensity for said first exon in said second sample; and
(e) identifying said first exon as being differentially spliced if the absolute value of the splicing index obtained in (d) is greater than a threshold value. - View Dependent Claims (2, 3, 4, 6, 7, 8, 9)
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5. The method of claim 5 wherein a first value of ni,j,k is calculated for a first sample and a second value of ni,j,k is calculated for a second sample and the splicing index is calculated by dividing the value of the ni,j,k for said first sample by the ni,j,k for said second sample to obtain a third value and calculating the log2 of said third value.
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10. A computer-implemented method for identifying exons that are differentially spliced between a first and second sample comprising:
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a. obtaining an exon intensity measurement for a first exon in a first gene in said first sample and in said second sample;
b. obtaining a gene level measurement for said first gene in said first sample and in said second sample;
c. obtaining a first normalized intensity measurement for said first exon in said first sample and a second normalized intensity measurement for said first exon in said second sample using the following equation;
ni,j,k=ei,j,k/gi,k where ei,j,k is the exon signal estimate of the i exon, j experiment, and k gene and gi,k is the gene level signal estimate of the j experiment and k gene;
d. obtaining a splicing index measurement for said first exon, wherein a first value of ni,j,k is calculated for a first sample and a second value of ni,j,k is calculated for a second sample and the splicing index is calculated by dividing the value of the ni,j,k for said first sample by the ni,j,k for said second sample to obtain a third value and calculating the log2 of said third value; and
e. identifying said first exon as being differentially spliced if the absolute value of the splicing index obtained in (d) is greater than a threshold value. - View Dependent Claims (11, 12)
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13. A method for predicting if an exon in a gene is differentially spliced in a first and a second sample sets, comprising:
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obtaining a first exon level intensity measurement for said exon in said first sample set and a second exon level intensity measurement for said exon in said second sample set;
assume under the null hypothesis of no alternative splicing for said exon that all exons of said gene are expressed at levels that are proportional to each other in said first and second sample sets;
fit a model that predicts exon response under said null hypothesis;
construct a statistic that measures how deviant the observed data is from the model;
use the statistic to construct a p-value; and
analyze the p-value to predict if the exon is differentially spliced. - View Dependent Claims (14, 15)
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Specification