Table 3 Description of primers that were used in the present work for genome-based deletion of the catB gene (NCgl2318) and integration of the tuf-promoter for overexpression of the catA gene (NCgl2319) in Corynebacterium glutamicum
Name | Sequence | Use | AT [°C] |
|---|---|---|---|
ΔcatB TS1 FW | 5′-TTAACAATTGGGATCCTCTAGACCCACTCCAGAGCATTGGGGTGTTT-3′ | Amplification of TS1; TS1 FW+ overlap to pClik, TS 1 RV+ overlap to TS2 FW | 55 |
ΔcatB TS1 RV | 5′-AGGTCCAATCGGCACCGCAGCAT-3′ | ||
ΔcatB TS2 FW | 5′-GGATGCTGCGGTGCCGATTGGACCCTCAAGCGCCTTCATGGTCTCAAC-3′ | Amplification of TS2; TS2 FW+ overlap to TS1 RV, TS 2 RV+ overlap to pClik | 57 |
ΔcatB TS2 RV | 5′-GCAGCCCGCTAGCGATTTAAATCCCTGGGCACCCATGGCGCCCAG-3′ | ||
Pef-tu-catA-TS1 FW | 5′-AATTGGGATCCTCTAGACCCACGATTCCCTCTTCAGGGCG-3′ | Amplification of TS1; TS1 FW+ overlap to pClik, TS1 RV+ overlap to Pef-tu | 61 |
Pef-tu-catA-TS1 RV | 5′-CATTCGCAGGGTAACGGCCAGTCATACTCCTAGAGACTTTGGGGG-3′ | ||
Pef-tu-catA-Pef–tu FW | 5′-AAAGTCTCTAGGAGTATGACTGGCCGTTACCCTGCGAATG-3′ | Amplification of Pef–tu; Pef–tu FW+ overlap to TS1 RV, Pef–tu RV+ overlap to TS2 FW | 62 |
Pef-tu-catA-Pef–tu RV | 5′-ATCTGTTCAGCTGAAGTCATTGTATGTCCTCCTGGACTTCGTGG-3′ | ||
Pef-tu-catA-TS2 FW | 5′-GAAGTCCAGGAGGACATACAATGACTTCAGCTGAACAGATCG-3′ | Amplification of TS2; TS2 FW+ overlap to Pef-tu, TS2 RV + overlap to pClik | 57 |
Pef-tu-catA-TS2 RV | 5′-CCGCTAGCGATTTAAATCCCTAGTATCCGTCCTCATCTGCG-3′ | ||
PR154 | 5′-ATTGTCTGTTGTGCCCAGTCATAG-3′ | Validation of vector pClik int sacB (including respective insert) | 53 |
PR318 | 5′-AATAATAGTGAACGGCAGGT-3′ |