Simultaneous quantification of alternatively spliced transcripts in a single droplet digital PCR reaction

Simultaneous quantification of alternatively spliced transcripts in a single droplet digital PCR reaction

Human telomerase reverse transcriptase (hTERT) is an essential component required for telomerase activity and telomere maintenance. Several alternatively spliced forms of hTERT mRNA have been reported in human primary and tumor cells. Currently, however, there is no sensitive and accurate method for...

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Veröffentlicht in:BioTechniques. - 1993. - 56(2014), 6 vom: 13. Juni, Seite 319-25
1. Verfasser: Sun, Bing (VerfasserIn)
Weitere Verfasser: Tao, Lian, Zheng, Yun-Ling
Format: Online-Aufsatz
Sprache:English
Veröffentlicht: 2014
Zugriff auf das übergeordnete Werk:BioTechniques
Schlagworte:Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't RNA alternative splicing ddPCR droplet digital PCR hTERT qPCR real-time PCR telomerase mehr... telomere DNA Primers DNA-Binding Proteins RNA, Messenger TERT protein, human EC 2.7.7.49 Telomerase
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520 |a Human telomerase reverse transcriptase (hTERT) is an essential component required for telomerase activity and telomere maintenance. Several alternatively spliced forms of hTERT mRNA have been reported in human primary and tumor cells. Currently, however, there is no sensitive and accurate method for the simultaneous quantification of multiple alternatively spliced RNA transcripts, such as in the case of hTERT. Here we show droplet digital PCR (ddPCR) provides sensitive, simultaneous digital quantification in a single reaction of two alternatively spliced single deletion hTERT transcripts (α-/β+ and α+/β-) as well as the opportunity to manually quantify non-deletion (α+/β+) and double deletion (α-/β-) transcripts. Our ddPCR method enables direct comparison among four alternatively spliced mRNAs without the need for internal standards or multiple primer pairs specific for each variant as real-time PCR (qPCR) requires, thus eliminating potential variation due to differences in PCR amplification efficiency 
650 4 |a Journal Article 
650 4 |a Research Support, N.I.H., Extramural 
650 4 |a Research Support, Non-U.S. Gov't 
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650 7 |a Telomerase  |2 NLM 
650 7 |a EC 2.7.7.49  |2 NLM 
700 1 |a Tao, Lian  |e verfasserin  |4 aut 
700 1 |a Zheng, Yun-Ling  |e verfasserin  |4 aut 
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