Restriction digests of the clone give the following sizes (kb): BglII--4.5; EcoRI--4.5; SalI/BglII--3.5, 1.0. The marker, promoter and 3HA tag are flanked by the F4 primer site (5') and the R3 primer site (3'). The size of the F4/R3 PCR product is 1602 nt. F4 primer sequence: 5'-(gene-specific sequence)GAATTCGAGCTCGTTTAAAC-3'. R3 primer sequence: 5'(gene-specific sequence)GCACTGAGCAGCGTAATCTG-3'. The gene-specific sequence of these primers are chosen to correspond to the reverse complement of the N-terminal codons of the target gene. The sequences shown here are the reverse 3'. Contains a functional truncated (700 nt) ADH promoter. The promoter is not down regulated during the exponential phase. |
