Before PIC formation, three complexes, DNA-TFIIA-TFIIB-TBP, Pol II-TFIIF and TFIIH-TFIIE were incubated as three separate samples at room temperature for 5-10 min. Then, the pre-assembled samples were mixed and further incubated at 30°C for 1 h. Transcription was initiated by the addition...
Affinity-purified anti-Pat1 antibody was diluted 1:1000 in TBS-T plus 5% dry milk and incubated with the membrane overnight at 4 °C. Anti-RickA101 was used as a loading control by diluting serum 1:2,000 in TBS-T plus 5% dry milk and incubating at room temperature for 1 h. ...
To tether HGN to the exterior of the liposome membrane, HGN are mixed with liposomes with thiol-PEG in the membrane overnight at room temperature at relative concentrations to provide 1–10 HGN per liposome. A single HGN on a liposome is sufficient for generating nanobubbles and lysing the lip...