Restriction digests of the clone give the following sizes (kb): BglII--5.6; EcoRI--5.6; BglII/EcoRI--4.4, 1.3. The marker, promoter and GST cassette is flanked by the F4 primer site (5') and the R4 primer site (3'). The size of the F4/R4 PCR product is 2692 nt. F4 primer sequence: 5'-(gene-specific sequence)GAATTCGAGCTCGTTTAAAC-3'. R6 primer sequence: 5'-(gene-specific sequence) CATTGTATATGAGATAGTTGA-3'. R4 primer sequence: 5'-(gene-specific sequence) ACGCGGAACCAGATCCGATT-3'. The gene-specific sequences of these primers are chosen to correspond to the reverse complement of the N-terminal codons of the target gene. Contains a functional truncated (700 nt) ADH promoter. The promoter is not down regulated during the exponential phase. |
