ReViTA: A novel in vitro transcription system to study gene regulation

ReViTA (Reverse in Vitro Transcription Assay) is a novel in vitro transcription-based method to study gene expression under the regulation of specific transcription factors. The ReViTA system uses a plasmid with a control sequence, the promoter region of the studied gene, the transcription factor of interest, and an RNA polymerase saturated with sigma 70. The main objective of this study was to evaluate the method; thus, as a proof of concept, two different transcription factors were used, a transcriptional inducer, AlgR, and a repressor, LexA, from Pseudomonas aeruginosa. After the promoters were incubated with the transcription factors, the plasmid was transcribed into RNA and reverse transcribed to cDNA. Gene expression was measured using qRT-PCR. Using the ReViTA plasmid, transcription induction of 55% was observed when AlgR protein was added and a 27% transcription reduction with the repressor LexA, compared with the samples without transcription factors. The results demonstrated the correct functioning of ReViTA as a novel method to study transcription factors and gene expression. Thus, ReViTA could be a rapid and accessible in vitro method to evaluate genes and regulators of various species.

Automated summary

ReViTA (Reverse in Vitro Transcription Assay) is a novel in vitro transcription-based method used to study gene expression under specific transcription factors. The method was evaluated using two different transcription factors, AlgR and LexA, and demonstrated successful gene expression regulation. ReViTA has the potential to be a rapid and accessible method for studying genes and regulators in various species.