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Maximizing transcription of nucleic acids with efficient T7 promoters

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Item Type:Article
Title:Maximizing transcription of nucleic acids with efficient T7 promoters
Creators Name:Conrad, T. and Plumbom, I. and Alcobendas, M. and Vidal, R. and Sauer, S.
Abstract:In vitro transcription using T7 bacteriophage polymerase is widely used in molecular biology. Here, we use 5'RACE-Seq to screen a randomized initially transcribed region of the T7 promoter for cross-talk with transcriptional activity. We reveal that sequences from position +4 to +8 downstream of the transcription start site affect T7 promoter activity over a 5-fold range, and identify promoter variants with significantly enhanced transcriptional output that increase the yield of in vitro transcription reactions across a wide range of template concentrations. We furthermore introduce CEL-Seq(+) , which uses an optimized T7 promoter to amplify cDNA for single-cell RNA-Sequencing. CEL-Seq(+) facilitates scRNA-Seq library preparation, and substantially increases library complexity and the number of expressed genes detected per cell, highlighting a particular value of optimized T7 promoters in bioanalytical applications.
Keywords:AT Rich Sequence, Bacteriophage T7, Base Sequence, Genetic Promoter Regions, Genetic Templates, Genetic Transcription, Nucleic Acids, RNA-Seq, Single-Cell Analysis
Source:Communications Biology
Publisher:Springer Nature
Page Range:439
Date:14 August 2020
Official Publication:https://doi.org/10.1038/s42003-020-01167-x
PubMed:View item in PubMed

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