Revision as of 16:46, 23 September 2024

CMV-GraR-mRuby2

Usage and Biology

Contained within are two key genes: mRuby2, a red fluorescent protein for live-cell imaging, and graR, a regulator that might bind to PknB upon phosphorylating GraR. . GraR is known for its role in β-lactam resistance by upregulating cell wall biosynthesis genes, altering cell wall composition, and increasing expression of ABC-transporter (El-Halfawy et al., 2020),(Yang et al., 2012),(Meehl et al., 2007). The GraSR system is a two-component regulatory system that controls the expression of many genes involved in stress response, cell wall metabolism and virulence pathways. The GraSR system was found to control genes involved in stress response, cell wall metabolism and virulence pathways, in addition to playing an important role in CAMP resistance, thus greatly enhancing its importance as the main signal transduction pathway of Staphylococcus aureus (Falord et al., 2011). When activated by pknB, GraR binds to specific DNA sequences to regulate gene expression, in our case it presumably binds to a specific engineered promotor.

Cloning

This part was engineered with NEBBuilder® HIFI assembly method. First the backbone was linearized with NheI and BamHI and matching ends of gene and backbone ensured seamless cloning of GraR. In mammalian systems, this part is useful for studying the potential interactions between GraR and PknB. Sequence and Features