Open AccessCombining protein and RNA quantification to evaluate promoter activity by using dual-color fluorescent reporting systems
Author(s) -
Ying Peng,
Xin Huang,
Taizhong Huang,
Feng Du,
Xin Cui,
Zhuo Tang
Publication year - 2021
Publication title -
bioscience reports
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.938
H-Index - 77
eISSN - 1573-4935
pISSN - 0144-8463
DOI - 10.1042/bsr20211525
Subject(s) - mcherry , promoter , green fluorescent protein , transcription (linguistics) , ribozyme , rna , gene expression , gene , biology , microbiology and biotechnology , computational biology , genetics , linguistics , philosophy
Herein, Broccoli/mCherry and an EGFP/mCherry dual-color fluorescent reporting systems have been established to quantify the promoter activity at transcription and translation levels in eukaryotic cells. Based on those systems, four commonly used promoters (CMV and SV40 of Pol II and U6, H1 of Pol III) were accurately evaluated at both the transcriptional and translational levels by combining accurate protein and RNA quantification. Furthermore, we verified that Pol III promoters can induce proteins expression, and Pol II promoter can be applied to express RNA molecules with defined length by combining a self-cleaving ribozyme and an artificial poly(A) tail. The dual-color fluorescence reporting systems described here could play a significant role in evaluating other gene expression regulators for gene therapy.