Restriction digests of the clone give the following sizes (kb): BamHI--7.2; EcoRI--4.8, 1.4, 0.94; PvuII--4.4, 2.7. The insert contains the following restriction sites (approximate kb from the 5' end): XbaI--0.72; EcoRI--0.99, 2.37; HindIII--1.70; ScaI--1.96. An SpeI digest removes an intact 1.334 kb MET15 cassette, that includes flanking sequences of 577 bp at the 5' end and 580 bp at the 3' end). MET15, MET17 and MET25 are synonymous. The sequence of the PCR-generated MET15 cassette has not been determined to eliminate the possibility of PCR error. It was selected based on the described functional assay. |
