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Transposon-encoded CRISPR-Cas systems direct RNA-guided DNA integration.
Klompe SE, Vo PLH, Halpin-Healy TS, Sternberg SH
Nature. 2019 Jul;571(7764):219-225. doi: 10.1038/s41586-019-1323-z. Epub 2019 Jun 12.
PubMed Article

Plasmids from Article

ID Plasmid Purpose  
130633pSL0283 (pTnsABC)Expresses V. cholerae CAST TnsA, TnsB, and TnsC from a T7 promoter.
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130634pSL0527 (pDonor)Encodes a mini-transposon derived from V. cholerae CAST system, with CmR cargo gene. Total transposon size = 977 bp.
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130635pSL0284 (pQCascade_entry)Expresses V. cholerae CAST TniQ, Cas8, Cas7, and Cas6 from one T7 promoter, and a CRISPR RNA from a second T7 promoter. The CRISPR array contains two BsaI sites for spacer cloning.
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130636pSL0762 (pQCascade_crRNA-nt)Expresses V. cholerae CAST TniQ, Cas8, Cas7, and Cas6 from one T7 promoter, and a CRISPR RNA from a second T7 promoter. The CRISPR array encodes a non-targeting guide RNA.
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130637pSL0828 (pQCascade_crRNA-4)Expresses V. cholerae CAST TniQ, Cas8, Cas7, and Cas6 from one T7 promoter, and a CRISPR RNA from a second T7 promoter. The CRISPR array encodes a guide RNA that targets lacZ.
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130638pSL0373 (His-MBP-TniQ)Expresses V. cholerae CAST TniQ from a T7 promoter, with N-terminal His10-MBP-TEVsite tag for purification.
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130639pSL0374 (His-MBP-Cas8_Cas7_Cas6)Expresses V. cholerae CAST Cas8, Cas7, and Cas6 from a T7 promoter, with N-terminal His10-MBP-TEVsite tag on Cas8 for purification.
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130640pSL0375 (His-MBP-TniQ_Cas8_Cas7_Cas6)Expresses V. cholerae CAST TniQ, Cas8, Cas7, and Cas6 from a T7 promoter, with N-terminal His10-MBP-TEVsite tag on TniQ for purification.
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130641pSL0376 (His-MBP-StrepII-TniQ)Expresses V. cholerae CAST TniQ from a T7 promoter, with N-terminal His10-MBP-TEVsite-StrepII for purification.
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130642pSL0426 (CRISPR with spc3)Expresses CRISPR RNA from a T7 promoter for VchCAST system. The CRISPR array encodes a guide RNA with spacer 3 from the native CRISPR.
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130643pSL0915 (CRISPR with target 3)Expresses CRISPR RNA from a T7 promoter for VchCAST system. The CRISPR array encodes a guide RNA that targets lacZ.
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130644pSL0715 (pDonor_R1-57)Encodes a mini-transposon derived from V. cholerae Tn6677 CAST, with CmR cargo gene and shortened right end. Total transposon size = 910 bp.
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130645pSL0721 (pDonor_L1-105)Encodes a mini-transposon derived from V. cholerae Tn6677 CAST, with CmR cargo gene and shortened left end. Total transposon size = 933 bp.
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130646pSL0746 (pDonor_R-MmeI)Encodes a mini-transposon derived from V. cholerae Tn6677 CAST, with CmR cargo gene. The mutated right end introduces an MmeI recognition site (used for Tn-seq). Total transposon size = 977 bp.
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130647pSL0747 (pDonor_L-MmeI)Encodes a mini-transposon derived from V. cholerae Tn6677 CAST, with CmR cargo gene. The mutated left end introduces an MmeI recognition site (used for Tn-seq). Total transposon size = 977 bp.
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Antibodies from Article