A Step-by-Step Guide for the Production of Recombinant Fluorescent TAT-HA-Tagged Proteins and their Transduction into Mammalian Cells.

Investigating the function of target proteins for functional prospection or therapeutic applications typically requires the production and purification of recombinant proteins. The fusion of these proteins with tag peptides and fluorescently derived proteins allows the monitoring of candidate proteins using SDS-PAGE coupled with western blotting and fluorescent microscopy, respectively. However, protein engineering poses a significant challenge for many researchers.

Clarin-2 gene supplementation durably preserves hearing in a model of progressive hearing loss.

Hearing loss is a major health concern affecting millions of people worldwide with currently limited treatment options. In clarin-2-deficient Clrn2 mice, used here as a model of progressive hearing loss, we report synaptic auditory abnormalities in addition to the previously demonstrated defects of hair bundle structure and mechanoelectrical transduction. We sought an in-depth evaluation of viral-mediated gene delivery as a therapy for these hearing-impaired mice.

When translation elongation is impaired, the mRNA is uniformly destabilized by the RNA degradosome, while the concentration of mRNA is altered along the molecule.

mRNA sits at the crossroads of transcription, translation and mRNA degradation. Many questions remain about the coupling of these three processes in Escherichia coli and, in particular, how translation may have an effect on mRNA degradation and transcription. To characterize the interplay between mRNA degradation and translation while accounting for transcription, we altered the translation initiation or elongation and measured the effects on mRNA stability and concentration.

5'UTR sequences influence protein levels in by regulating translation initiation and mRNA stability.

A set of 41 synthetic 5'UTRs with different theoretical translation initiation rates were generated to explore the role of 5'UTRs in the regulation of protein levels in . The roles of the synthetic 5'UTRs in regulating the expression of different reporter genes were analyzed . Protein levels varied substantially between the different constructs but for most of the 5'UTRs, protein levels were not correlated with theoretical translation initiation rates.