Restriction digests of the clone give the following sizes (kb): SalI--3.35; SacI/KpnI--2.8, 0.5; XhoI--3.1, 0.25. Convenient cloning vector allowing generation of a fusion protein with the DNA-binding domain of GAL4 (aa 3-147). Multiple cloning sites present at both ends of the GAL4 segment permit in-frame fusions with the DNA-binding domain in the middle or at the C-terminus of the fusion product. pBR322 was modified by deletion of bp 75-2352, destruction (by filling in) of the HindIII site, deletion of a 100 bp AatII/ClaI fragment (nt 4285 and 22 of pBR322), and inserting an XbaI linker into the latter deletion junction. Transcription from the T3 promoter gives the sense strand. The order of the major features of this plasmid is: pMB1 ori - lacI (non-functional) - lacZ'/ - T3 promoter - SacI/MCS/SpeI - GAL4 - ClaI/MCS/KpnI - T7 promoter - /lacZ' - f1 ori - ampR. |
