Difference between revisions of "Part:BBa J176013"

Revision as of 02:36, 22 November 2011

VP64

Tetrameric VP16 transcription activator domain

Usage and Biology

• Chassis: mammalian cells
• Mammalian expression vector required
• Protein domain; requires promoter, start codon, stop codon, and polyA signal for proper expression
• VP64 doesn't do much as a free-floating domain. Fuse it to a DNA binding domain
  

VP64 is a transcriptional activator composed of four tandem copies of VP16 (Herpes Simplex Viral Protein 16, amino acids 437-447, DALDDFDLDML) connected with glycine-serine linkers. When fused to another protein domain that can bind near the promoter of a gene, VP64 acts as a strong transcriptional activator. This module is a classic molecular biology tool, which pre-dates fancy terms like "synthetic biology." See the References section for some history on VP16 and VP64.

Characterization

VP64 activity is sensitive to promoter proximity.
In our hands, VP64 activity is maximized when it is targeted to DNA sequences that are ~40-80 base pairs away from a minimal promoter. When fused to the Gal4 DNA binding domain (BBa_J176020) and targeted it to Gal (UAS) DNA elements (BBa_J176019) that are assembled immediately adjacent to the HSVtkTATA minimal promoter (BBa_J176011), we observe weak activation, and perhaps repression, of a fluorescent reporter gene. In contrast, when a small spacer (BBa_J176039) is inserted between the Gal4 binding sites and the promoter, reporter gene activity greatly increases.

(Data will be posted soon)

Sequence and Features