Restriction digests of the clone give the following sizes (kb): BamHI--7.3, 2.8; EcoRI--4.4, 3.9, 1.2, 0.5; HindIII--4.6, 2.2, 1.7, 0.9, 0.3. The insert contains the following restriction sites (approximate nt from the 5' end): BglI--0.4, 0.6; HindIII--1.0, 1.36, 2.24, 2.4; PstI--1.6, 1.64; EcoRI--2.3. Extends from 24 nt upstream of the major initiation codon to 360 nt downstream of the termination codon and thus encodes a protein of 795 amino acids. pART1 was constructed by inserting a 1.2 kb Schizosaccharomyces pombe ars fragment into the EcoRI site of pUC118 and a 2.2 kb fragment containing Saccharomyces cerevisiae LEU2 into the HindIII site. A 700 bp SphI/PstI fragment containing the adh promoter of Sc. pombe was inserted into polylinker sites. Transcription is driven from the SphI site toward the remaining PstI, SalI and BamHI sites through the insert sequence. |
