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Wang H, Wang H, Duan X, et al. Highly sensitive and multiplexed quantification of mRNA splice variants by the direct ligation of DNA probes at the exon junction and universal PCR amplification. Chem Sci. 2017;8(5):3635-3640doi: 10.1039/c7sc00094d.
Wang, H., Wang, H., Duan, X., Sun, Y., Wang, X., & Li, Z. (2017). Highly sensitive and multiplexed quantification of mRNA splice variants by the direct ligation of DNA probes at the exon junction and universal PCR amplification. Chemical science, 8(5), 3635-3640. https://doi.org/10.1039/c7sc00094d
Wang, Honghong, et al. "Highly sensitive and multiplexed quantification of mRNA splice variants by the direct ligation of DNA probes at the exon junction and universal PCR amplification." Chemical science vol. 8,5 (2017): 3635-3640. doi: https://doi.org/10.1039/c7sc00094d
Wang H, Wang H, Duan X, Sun Y, Wang X, Li Z. Highly sensitive and multiplexed quantification of mRNA splice variants by the direct ligation of DNA probes at the exon junction and universal PCR amplification. Chem Sci. 2017 May 01;8(5):3635-3640. doi: 10.1039/c7sc00094d. Epub 2017 Mar 01. PMID: 28580102; PMCID: PMC5437374.
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Chem Sci. 2017 May 01;8(5):3635-3640. doi: 10.1039/c7sc00094d. Epub 2017 Mar 01.

Highly sensitive and multiplexed quantification of mRNA splice variants by the direct ligation of DNA probes at the exon junction and universal PCR amplification.

Chemical science

Honghong Wang, Hui Wang, Xinrui Duan, Yuanyuan Sun, Xiangdong Wang, Zhengping Li

Affiliations

  1. Key Laboratory of Analytical Chemistry for Life Science of Shaanxi Province , School of Chemistry and Chemical Engineering , Shaanxi Normal University , Xi'an 710062 , Shaanxi Province , P. R. China . Email: [email protected] ; Email: [email protected].

PMID: 28580102 PMCID: PMC5437374 DOI: 10.1039/c7sc00094d

Abstract

Alternative messenger RNA (mRNA) splicing is a basic mechanism of gene regulation. In general, reverse transcription and polymerase based primer extension limit the sensitivity and selectivity of the current detection of mRNA splice variants, respectively. Here, we show that, using the ligation of two properly designed probes at the exon junction combined with universal PCR amplification, as little as a single copy of a mRNA splice variant per cell can be accurately determined, and the dynamic range covers six orders of magnitude. Three mRNA splice variants were measured from total RNA samples derived from different cell lines. Moreover, by encoding the ligation probes with different lengths, multiplexed mRNA splice variants can be simultaneously detected in one-tube PCR amplification using electrophoretic separation.

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