Accelrys ds gene software

We designed the following oligonucleotides and plasmids according to the method of Koibuchi et al.^{29 (link)} F2 (chick lysozyme TRE at nucleotides −2358 to −2326; half-sites arranged as an inverted palindrome with nucleotide gap of 6 (ref. 35 (link))); palindromic (typical TREs, which designed not to contain a putative RORE); DR4-1 (DR4 sequence containing AT-rich sequences at the 5′ end of the upstream half-site to serve as a putative RORE); and DR4-2 (DR4 oligonucleotide lacking an AT-rich sequence).^{36 (link)} These oligonucleotides were cloned in the psi-CHECK2 vector (Thermo Fisher Scientific, USA) containing a viral thymidine kinase promoter coupled to the firefly luciferase-labelled gene which was used as the internal reference to remove the transfection efficiency differences between groups, and the Renilla luciferase reporter gene.^{37 (link)} These reporter plasmids were sequenced to ensure that only a single copy of the TRE had been incorporated (Fig. 2A).
Accelrys DS Gene software (Accelrys Software, Inc., U.S.A.) was adopted to analyze the enzyme-cutting site of the psiCHECK2 promoter between the Poly A and hsv-tk promoter. The Bbs I enzyme was selected and the cutting site is shown in Fig. 2B. The nucleotide sequences of the double-stranded oligonucleotides containing TRE or RORE are shown in Fig. 2C.