4/14/2023 0 Comments Srdx transcriptionNatural TALEs are transcription factors used by plant-pathogenic bacteria in the genus Xanthomonas. Transciption activator-like effector (TALE) technology has recently emerged as an alternative robust and efficient genome editing tool. However, the DNA-binding domain of zinc-finger nucleases are challenging to design and require experimental optimization, and the target sequences of meganucleases are limited. Zinc-finger nucleases (ZFNs) and meganucleases have been well-established as tools to edit specific sites in complex genomes. This system offers a robust and easy-to- use tool for real-time monitoring and quantifying gene expression in mammalian cells. We have designed, constructed and validated a novel dual reporter system for assessing TALE mediated gene regulations. We validated both sensitivity and specificity of this dual-reporter system in mammalian cells, and demonstrated that this dual reporter system is robust and potentially amenable to high throughput (HTP) applications. We established a dual reporter system that was specifically designed for real-time monitoring and quantifying gene expression mediated by TALEs. Although the efficient construction of TALEs has been established, robust functional tools to assess their functions remain lacking. The modularity of TALEs DNA binding domain enables sequence-specific perturbation and offers broad applications in genetic and epigenetic studies. These results suggest that genetic manipulation of AtMYī49 may provide a novel way to improve salt tolerance in plants.Transcription activator-like effectors (TALEs) are a class of naturally occurring transcription effectors that recognize specific DNA sequences and modulate gene expression. Additionally, increased AtMYB49 expression elevated Ca²⁺ level in leaves and improved antioxidant capacity by up‐regulating genes encoding peroxidases and late embryogenesis abundant proteins. Importantly, cuticular transpiration, chlorophyll leaching and toluidine blue‐staining assays revealed a link between increased AtMYB49‐mediated cutin deposition in leaves and enhanced salt tolerance. Biochemical analysis indicated that AtMYB49 modulated cutin deposition in the leaves. Some of these differentially expressed genes, including MYB41, ASFT, FACT and CYP86B1, were also shown to be the direct targets of AtMYB49 and activated by AtMYB49. Transcriptome analysis revealed that many genes belonging to the category “cutin, suberin and wax biosyntheses” were markedly up‐regulated and down‐regulated in OX49 and SRDX49 plants, respectively, under normal and/or salt stress conditions. Characterization of loss‐of‐function myb49 mutants, and chimeric AtMYB49‐SRDX‐overexpressing SRDX49 transcriptional repressor and AtMYB49‐overexpressing (OX49) overexpressor plants demonstrated a positive role of AtMYB49 in salt tolerance. Here, we showed that the transcriptional activation of cutin deposition and antioxidant defence by the R2R3‐type MYB transcription factor AtMYB49 contributed to salt tolerance in Arabidopsis thaliana. Salt stress activates defence responses in plants, including changes in leaf surface structure. These results suggest that genetic manipulation of AtMYB49 may provide a novel way to improve salt tolerance in plants.
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