A broad-host-range CRISPRi toolkit for silencing gene expression in Burkholderia.
Hogan AM, Rahman ASMZ, Lightly TJ, Cardona ST
ACS Synth Biol. 2019 Sep 6. doi: 10.1021/acssynbio.9b00232.
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Plasmids from Article
| ID | Plasmid | Purpose |
|---|---|---|
| 129389 | pAH25-SceI | Derived from pDAI-SceI. Tetracycline resistance removed and replaced with chloramphenicol resistance cassette from pKD3. |
| 129390 | pAH-CTX1-rha | Derived from mini-CTX1. An integrase vector that can integrate the rhamnose-inducible promoter system into the chromosome of a host bacterium. |
| 129391 | pAH-CTX1-rhadCas9 | Derived from pAH-CTX1-rha. The Burkholderia cenocepacia codon-optimized dCas9 cloned downstream of PrhaBAD. Hence, dCas9 is controlled by the rhamnose-inducible promoter system. |
| 129392 | pAH-CTX1-rhadCas9-native | Derived from pAH-CTX1-rha. The native Streptococcus pyogenes dCas9 gene cloned downstream of PrhaBAD. Non-codon-optimized dCas9 version of pAH-CTX1-rhadCas9. |
| 129463 | pSCB2-sgRNA | Template plasmid for guide sequence insertion via inverse PCR. Derived from pSCrhaB2-gRNA (see paper). |
| 129464 | pgRNA-guideless | Derived from pSCB2-sgRNA. 20 nucleotide binding sequence of sgRNA removed by inverse PCR. Used as a negative control. |
| 129465 | pgRNA-non-target | Derived from pSCB2-sgRNA. 20 nucleotide sequence added as sgRNA binding region by inverse PCR. Used as a negative control. |
| 154397 | pAH18 | Derived from pGPI-SceI; Trimethoprim resistant; carries I-SceI gene; contains 475 bp from both upstream and downstream flanking regions between the FRT sites on pAHCTX1-rhadCas9 |