| Strain |
Relevant characteristics |
Source or reference |
|
Escherichia coli DH5α |
F-
endA1 glnV44 thi-1 recA1 relA1 gyrA96 deoR nupG φ80dlacZΔM15 Δ(lacZYA-argF)U169, hsdR17(rK-mK+), λ-
|
Lab stock |
|
Escherichia coli ER1821 |
F-
endA1 glnV44 thi-1 relA1? e14-(mcrA-) rfbD1? spoT1? Δ(mcrC-mrr)114::IS10
|
Lab stock; New England Biolabs |
|
Clostridium pasteurianum ATCC 6013 |
Wild-type |
American Type Culture Collection |
|
Clostridium pasteurianum ∆cpaAIR
|
Markerless cpaAIR deletion mutant |
This study |
| Plasmid |
Relevant characteristics |
Source or reference |
| pFnuDIIMKn |
M.FnuDII methyltransferase plasmid for methylation of E. coli-C. pasteurianum shuttle vectors (KmR; p15A ori) |
49 |
| pMTL83151 |
E. coli-Clostridium shuttle vector (CmR; ColE1 ori; pCB102 ori) |
57 |
| pMTL85141 |
E. coli-Clostridium shuttle vector (CmR; ColE1 ori; pIM13 ori) |
57 |
| pCas9 |
E. coli cas9 and tracrRNA expression vector (CmR; p15A ori) |
27 |
| pCas9gRNA-cpaAIR |
Type II CRISPR expression vector containing cas9 and gRNA targeted to the C. pasteurianum cpaAIR gene |
This study |
| pCas9gRNA-delcpaAIR |
Type II CRISPR genome editing vector derived by inserting a cpaAIR deletion editing cassette into pCas9gRNA-cpaAIR |
This study |
| p85Cas9 |
cas9 expression vector derived by inserting cas9 with its native promoter from pCas9 into pMTL85141 |
This study |
| p83Cas9 |
cas9 expression vector derived by inserting cas9 and the tracrRNA from pCas9 into pMTL83151 |
This study |
| p85delCas9 |
Derived by deleting the cas9 promoter from p85cas9 |
This study |
| pSpacer18 |
C. pasteurianum protospacer 18 construct lacking flanking sequences |
This study |
| pSpacer18-5′ |
C. pasteurianum protospacer 18 construct including 5′ protospacer-adjacent sequence |
This study |
| pSpacer18-3′ |
C. pasteurianum protospacer 18 construct including 3′ protospacer-adjacent sequence |
This study |
| pSpacer18-flank |
C. pasteurianum protospacer 18 construct including flanking protospacer-adjacent sequence |
This study |
| pSpacer24 |
C. pasteurianum protospacer 24 construct lacking flanking sequences |
This study |
| pSpacer24-5′ |
C. pasteurianum protospacer 24 construct including 5′ protospacer-adjacent sequence |
This study |
| pSpacer24-3′ |
C. pasteurianum protospacer 24 construct including 3′ protospacer-adjacent sequence |
This study |
| pSpacer24-flank |
C. pasteurianum protospacer 24 construct including flanking protospacer-adjacent sequence |
This study |
| pSpacer30 |
C. pasteurianum protospacer 30 construct lacking flanking sequences |
This study |
| pSpacer30-5′ |
C. pasteurianum protospacer 30 construct including 5′ protospacer-adjacent sequence |
This study |
| pSpacer30-3′ |
C. pasteurianum protospacer 30 construct including 3′ protospacer-adjacent sequence |
This study |
| pSpacer30-flank |
C. pasteurianum protospacer 30 construct including flanking protospacer-adjacent sequence |
This study |
| pCParray-cpaAIR |
Type I-B CRISPR expression vector containing a synthetic CRISPR array targeted to the C. pasteurianum cpaAIR gene |
This study |
| pCParray-delcpaAIR |
Type I-B CRISPR genome editing vector derived by inserting a cpaAIR deletion editing cassette into pCParray-cpaAIR |
This study |
