The pET-49b(+) vector carries N-terminal GST•Tag™ and His•Tag? coding sequences followed by a recognition site for the human rhinovirus (HRV) 3C protease. This protease is highly specifi c for cleavage of the sequence LEVLFQ↓GP , and is active at low temperatures . pET-49b(+) also contains an optional C-terminal thrombin recognition site followed by an S•Tag™ coding sequence. Unique restriction .sites are shown on the circle map. Note that the sequence is numbered by the pBR322 convention, so the T7 expression region is reversed on the circle map. The cloning/expression region of the coding strand transcribed by T7 RNA polymerase is shown below. The f1 origin is oriented so that infection with the helper phage will produce virions containing single-stranded DNA that corresponds to the coding strand. Therefore, single-stranded sequencing should be performed using the AS S•Tag 18mer Primer (Cat. No. 71262-3).
