Table 3 Strains and plasmids used in this study

Strains

E. coli DH5α

F⁻ ϕ80lacZΔM15 Δ(lacZYAargF)U169 endA1 recA1 hsdR17 (r_K⁻ m_K⁺) supE44 λ⁻thi-1 gyrA96 relA1 phoA

[86]

E. coli S17-1 λpir

F⁻thi pro hsdR hsdM⁺recA RP4-2-Tc::Mu- Km::Tn7 λpir Tp^R Sm^R

[87]

V. natriegens

Wild-type strain DSM 759 (ATCC 14048), referred to as WT

German Collection of Microorganisms and Cell Cultures

V. natriegens ΔligK

Deletion of ligK (PN96_18530) in V. natriegens WT

This study

V. natriegens Δvnp12

Deletions of VNP regions 1 (PN96_04290 to PN96_04520) and 2 (PN96_06880 to PN96_07090) in V. natriegens WT

[59]

V. natriegens Δvnp12 ΔaceE

Deletion of aceE (PN96_01335) in V. natriegens Δvnp12

This study

V. natriegens Δvnp12 ΔaceE Δdns::katG_Ec-1xFLAG

Replacing dns locus (PN96_00865) in V. natriegens Δvnp12 ΔaceE with katG from E. coli DH5α combined with a 1xFLAG tag

This study

V. natriegens Δvnp12 ΔaceE ΔligK

Deletion of ligK (PN96_18530) in V. natriegens Δvnp12 ΔaceE

This study

C. glutamicum

Wild-type strain ATCC 13,032

[88]

C. glutamicum ΔaceE Δpqo ΔldhA ΔC–T ilvN ΔalaT ΔavtA

Engineered ATCC 13,032 for the production of pyruvate

[19]

Plasmids

pDM4

oriV_R6K, oriT_RP4, sacB, Cm^R

[89]

pDM4-ΔaceE

pDM4 plasmid carrying 500 bp homologous sequences upstream and downstream of aceE

This study

pST_116

ori_ColE1, tfoX, cas9, acrIIA4, sfgfp, gRNA scaffold, tetR, lacI, Cm^R

[63]

pST_116-ΔligK

pST_116 with sfgfp exchanged for gRNA spacer targeting ligK

This study

pST_116-Δdns

pST_116 with sfgfp exchanged for gRNA spacer targeting dns

This study

pET-22b(+)

ori_pBR322, Carb^R

Novagen

pJH001

ori_ColE1, Carb^R, 3 kb upstream and downstream flanks of dns, sfgfp

This study

pJH001- katG_Ec-1xFLAG

pJH001 with sfgfp exchanged for katG from E. coli DH5α combined with a 1xFLAG tag

This study

pEKEx2

oriV_pBL1(C.g.), oriV_ColE1(E.c), P_tac, lacI^q, Kan^R

[90]

pEKEx2-ligK

pEKEx2 with ligK under P_tac control

This study