Single well, single-common primer pair, dual probe, duplex qPCR assay for the quantification of mRNA splicing variants.

ABSTRACT: Quantifying the ratio of alternatively spliced mRNA variants of genes with known alternative splicing variants is highly relevant for many applications. Herein, we describe the validation of a quantitative PCR design for the simplified quantification of known mRNA splice variants. The assay uses a single-common primer pair, dual probe design for the determination of splicing variants in a single well configuration. We used murine XBP-1 splicing variants, XBP-1S and XBP-1U, to validate and demonstrate the performance characteristics of this approach. Using synthetic XBP-1S and XBP-1U cDNA as well as cDNA synthesized from mouse beta-cell line MIN6, we established the performance parameters and dynamic range of the assay. Reliable quantification of both variants at varying concentration gradi

SUBMITTER: Wang J 

PROVIDER: S-EPMC7903517 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature