Researchers shall have no right to modify or otherwise create variations of the nucleotide sequence of the HaloTag® gene. Researchers may use this product for research use only, no commercial use is allowed.
#HALO TAG SNAPGENE FULL#
If the researcher is not willing to accept the conditions of this limited use statement, and the product is unused, Promega will accept return of the unused product and provide the researcher with a full refund. When the donor and acceptor proteins interact, energy transfer occurs, resulting in increased NanoBRET™ signal.īY USE OF THIS PRODUCT, RESEARCHER AGREES TO BE BOUND BY THE TERMS OF THIS LIMITED USE STATEMENT. The HaloTag® fusions are labeled with the fluorescent HaloTag® NanoBRET™ 618 Ligand and serve as the energy acceptor. In these assays, the target protein fused to NanoLuc® luciferase or a subunit of NanoBiT® enzyme and serves as the bioluminescent BRET energy donor. NanoBRET™ technology is a proximity-based method dependent upon energy transfer from a luminescent donor target protein to a fluorescent acceptor. The HaloTag® fusion vectors are designed to be used as the fluorescent acceptor in NanoBRET™ assays that monitor interactions of a specific target protein along the UPS pathway. An emerging modality for small-molecule drug development is targeting proteins for degradation via the UPS with small-molecule degraders such as PROTACs or molecular glues. The majority of proteins and their abundance are regulated via the ubiquitin proteasome system (UPS), which uses ubiquitin conjugation to signal proteins that should be trafficked to the proteasome for degradation.
#HALO TAG SNAPGENE SOFTWARE#
The combined methods including the Gibson assembly, lentiviral mediated gene transfer, spinning disk-based live cell imaging, and software for quantification allow analyses of the endothelial cell junction dynamics under static and under shear stress conditions.Regulation of cellular protein homeostasis is critical for maintaining cell health and often altered by cellular treatments or disease states. Remodeling was accompanied by VE-cadherin plaque formation, indicating that this process is mediated by the formation of the actin-driven junction-associated intermittent lamellipodia, JAIL. Using such combinations, we found maintained cell junction integrity during shear stress-induced junction remodeling using VE-cadherin-EGFP.
Furthermore, classical fluorescence tags such as mCherry and EGFP were compared with self-labeling tags such as Halo and SNAP for their suitability to study junction dynamics in cultured endothelium, and found the self-labeling tags as useful tools. We used the Gibson assembly as a quick and cheap cloning system for introducing sequences into the lentiviral-based pFUGW vector. The development of viral vectors has contributed significantly to the genetic manipulation of endothelial cells. In this work, we describe advanced methods that allow investigating the dynamics of endothelial cell contacts. Advanced microscopic techniques, automation processing, and image analysis software was shown to improve the understanding of vascular biology. Endothelial cells of the vascular system are dynamic cells whose molecular adaptability is decisive for the adjustment of homeostasis and organ perfusion.
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