VP16 Tag

VP16 from herpes simple virus (HSV) is a strong transcriptional activator of immediate early viral genes.1 Due to its lack of DNA binding activity, VP16 is recruited to DNA via interaction with DNA binding proteins.2 Ptashne et al. showed that VP16 can function as a potent transcriptional activator in mammalian cells when fused to the DNA binding domain of the yeast transcriptional activator GAL4.3 These properties were taken into consideration in the development of a mammalian version of the Yeast Two-Hybrid System. The Yeast Two-Hybrid System is a powerful method for characterizing and screening for protein-protein interactions. In the original version of the system, GAL4 DNA binding domain is fused to protein X, and GAL4 activation domain is fused to protein Y. Neither hybrid is capable of activating transcription individually. Upon interaction between proteins X and Y, the DNA binding and activator domains are brought in close proximity. As a result, the transcriptional activity of GAL4 is reconstituted at the promoter, and monitored with a reporter gene. Additional two-hybrid systems were developed using different DNA binding and activation domains as well as different reporter genes.5-9 The VP16 activator is used in yeast and mammalian systems. In yeast, its DNA interaction partner is typically LexA.10 In mammalian systems, interaction between two proteins is detected through recruitment of VP16 and GAL4 DNA binding domain to GAL4 operators present in the reporter gene.7,9-11 Applications: Suitable for use in Western Blot and Immunoprecipitation. Other applications not tested. Recommended Dilution: Immunoprecipitation: 10ug/ml Western Blot: 1:1000 Optimal dilutions to be determined by the researcher. Storage and Stability: May be stored at 4 degrees C for short-term only. For long-term storage and to avoid repeated freezing and thawing, add sterile glycerol (40-50%), aliquot and store at -20 degrees C. Aliquots are stable for at least 6 months at -20 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.