科研之友
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摘要
Background: A sensitive, high-throughput method for monitoring pre-mRNA splicing on a genomic scale is needed to understand the spectrum of alternatively spliced mRNA in human cells.
Results: We adapted Molecular Inversion Probes (MIPs), a padlock-probe based technology, for the multiplexed capture and quantitation of individual splice events in human tissues. Individual MIP capture probes can be quantified using either DNA microarrays or high-throughput sequencing, which permits independent assessment of each spliced junction. Using our methodology we successfully identified 100% of our positive controls and showed that there is a strong correlation between the data from our alternative splicing MIP (asMIP) assay and quantitative PCR.
Conclusion: The asMIP assay provides a sensitive, accurate and multiplexed means for measuring pre-mRNA splicing. Fully optimized, we estimate that the assay could accommodate a throughput of greater than 20,000 splice junctions in a single reaction. This would represent a significant improvement over existing technologies.
- 出版日期2010-12-17
