SIV-rtTA acquires additional mutations in the U3 and TAR region upon long-term culturing. SIV-rtTA was cultured with dox in PM1 cells for up to 234 days. Cellular proviral DNA was isolated from 13 independent cultures at different times and the LTR region was subsequently PCR amplified and sequenced. The number of the culture (#) and the day of sampling are indicated on the left. The -90 to +130 U3/R region is shown with +1 indicating the transcription initiation site. The Sp1 and TATA box are shown in grey. The mutations that were introduced in TAR to abolish Tat-responsiveness are underlined. The additional nucleotide substitutions and deletions (Δ) observed in the SIV-rtTA cultures are indicated.