Restriction digests of the clone give the following sizes (kb): XbaI--4.6; NotI--3.0, 1.6. A 912 bp PCR product with HindIII and BsiWI ends and containing HO promoter sequences (-1720 to -1814 upstream from the ATG) was inserted into the HindIII-BsiWI digested pUC21-NotI. A 507 bp PCR product with EcoRI and SfiI ends and containing 3' end sequences of HO gene (1199 to 1699 downstream from the ATG) was inserted into the EcoRI-SfiI digested pUC21-NotI. This plasmid allows targeted integration at HO locus. |
