Restriction digests of the clone give the following sizes (kb): BamHI/EcoRI--3.0, 1.1; NcoI--4.1; PvuI--3.0, 1.1; PstI--4.1; PvuII--2.5, 0.9, 0.7. The insert contains the following restriction sites (kb from the 5' end): NcoI--0.365; PvuII--0.407; PstI--0.635; BspMI--0.911. The deletion removes codons 238 to 284 and introduces a frameshift resulting in a termination codon. The protein product is therefore 237 aa in length, including the bipartite DNA-binding domain. The EcoRI site at the 5' end and the BamHI site at the 3' end of the insert were generated by PCR. Transcription in vitro from the T7 RNA polymerase promoter gives the antisense strand. |
