Table 4.
Plasmids constructed in this work.
| Plasmid | Vector | Inserted Sequences 1 |
|---|---|---|
| FBA gene expression | ||
| pJJH2957 | pCXs22 | KlFBA1 with its flanking sequences cloned as an EcoRI/BamHI fragment |
| pJJH3018 | pCXs22 | ScFBA1 with its flanking sequences cloned as an EcoRI/BamHI fragment |
| pJJH3057 | pJJH2075 | Synthetic-yeast-optimized human ALDOA cloned as a BamHI/HindIII fragment under control of ScPFK2p |
| pJJH3069 | YEplac181 | ScPFK2p-ALDOA subcloned from pJJH3057 as an EcoRI/BamHI fragment |
| lacZ fusions | ||
| pJJH3041 | pJJH2031 | KlGDP1 promoter cloned as a HindIII/BamHI fragment |
| pJJH3042 | pJJH2031 | KlTDH2 promoter cloned as a HindIII/BamHI fragment |
| pJJH3043 | pJJH2031 | KlFBA1 promoter cloned as a HindIII/BamHI fragment |
| pJJH3061 | pJJH2031 | KlTDH1 promoter cloned as a HindIII/BamHI fragment |
1 All DNA fragments to be inserted were generated by PCR from genomic yeast DNA using oligonucleotides with the appropriate restriction sites. As an exception, the human ALDOA coding sequence was obtained by custom synthesis from GeneArt (ThermoFisher Scientific, Germany) with codons optimized for S. cerevisiae. All sequences were confirmed after cloning by custom Sanger sequencing (Seqlab, Göttingen, Germany) and the entire plasmid sequences and detailed descriptions of their constructions are available upon request.