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crispri plasmid backbone  (New England Biolabs)


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    Structured Review

    New England Biolabs crispri plasmid backbone
    Crispri Plasmid Backbone, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1647 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/crispri+plasmid+backbone/BsmBI-v2/pm41720075-1015-1-8
    Average 99 stars, based on 1647 article reviews
    crispri plasmid backbone - by Bioz Stars, 2026-10
    99/100 stars

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    Related Articles

    Plasmid Preparation:

    Article Title: Cyclic AMP is a critical mediator of intrinsic drug resistance and fatty acid metabolism in M. tuberculosis
    Article Snippet: Generation of individual CRISPRi and CRISPRi-resistant complementation strains Individual CRISPRi plasmids were cloned as previously described in Bosch et al., 2021 using Addgene plasmid #166886. .. Briefly, the CRISPRi plasmid backbone was digested with BsmBI- v2 (NEB #R0739L) and gel purified. sgRNAs were designed to target the non- template strand of the target gene ORF. .. For each individual sgRNA, two complementary oligonucleotides with appropriate sticky end overhangs were annealed and ligated (T4 ligase NEB # M0202M) into the BsmBI- digested plasmid backbone.

    Article Title: Compensatory evolution in NusG improves fitness of drug-resistant M. tuberculosis
    Article Snippet: Individual CRISPRi plasmids were cloned as described using Addgene plasmid 166886. .. In brief, the CRISPRi plasmid backbone was digested with BsmBI-v2 (NEB R0739L) and gel-purified. sgRNAs were designed to target the non-template strand of the target gene open reading frame (ORF). .. For each individual sgRNA, two complementary oligonucleotides with appropriate sticky end overhangs were annealed and ligated (T4 ligase NEB M0202 M) into the BsmBI-digested plasmid backbone.

    Article Title: The phenotypic landscape of the mycobacterial cell
    Article Snippet: .. Briefly, the CRISPRi plasmid backbone was digested with BsmBI-v2 (NEB #R0739L). .. For each individual sgRNA, two complementary oligonucleotides with appropriate overhangs were annealed and ligated (T4 ligase NEB # M0202M) into the BsmBI-v2 digested plasmid backbone.


    Article Title: Cell-free genomics reveals fundamental regulatory principles of the Mycobacterium tuberculosis transcription cycle.
    Article Snippet: Article Cell-free genomics reveals fundamental regulatory principles of the Mycobacterium tuberculosis transcription cycle

    Article Title: An activator of a two-component system controls cell separation and intrinsic drug resistance in Mycobacterium tuberculosis
    Article Snippet: Individual CRISPRi plasmids were cloned as described previously using Addgene plasmid 166886 [ ]. .. Briefly, the CRISPRi plasmid backbone was digested with BsmBI-v2 (NEB R0739L) and gel purified. sgRNAs were designed to target the non-template strand of the target gene ORF. .. For each individual sgRNA, two complementary oligonucleotides with appropriate sticky end overhangs were annealed and ligated (T4 ligase NEB M0202M) into the BsmBI-digested plasmid backbone.

    Article Title: Mapping unsolved lipidomes accelerates lipid discovery in major bacterial pathogens
    Article Snippet: Individual CRISPRi plasmids were generated following the protocol outlined in ( ) using Addgene plasmid #163631. .. Briefly, the CRISPRi plasmid backbone was digested with BsmBI-v2 (NEB #R0739L) and subsequently gel purified. sgRNAs were designed to target the non-template strand of the target gene ORF. .. For each sgRNA, two complementary oligonucleotides with compatible sticky-end overhangs were annealed and ligated into the BsmBI-digested plasmid backbone using T4 DNA ligase (NEB #M0202M).

    Article Title: Compensatory evolution in NusG improves fitness of drug-resistant M. tuberculosis.
    Article Snippet: Generation of individual CRISPRi strains Individual CRISPRi plasmids were cloned as described67 using Addgene plasmid 166886. .. In brief, the CRISPRi plasmid backbone was digested with BsmBI-v2 (NEB R0739L) and gel-purified. sgRNAs were designed to target the non-template strand of the target gene open reading frame (ORF). .. For each individual sgRNA, two complementary oligonucleotides with appropriate sticky end overhangs were annealed and ligated (T4 ligase NEB M0202 M) into the BsmBI-digested plasmid backbone.

    Purification:

    Article Title: Cyclic AMP is a critical mediator of intrinsic drug resistance and fatty acid metabolism in M. tuberculosis
    Article Snippet: Generation of individual CRISPRi and CRISPRi-resistant complementation strains Individual CRISPRi plasmids were cloned as previously described in Bosch et al., 2021 using Addgene plasmid #166886. .. Briefly, the CRISPRi plasmid backbone was digested with BsmBI- v2 (NEB #R0739L) and gel purified. sgRNAs were designed to target the non- template strand of the target gene ORF. .. For each individual sgRNA, two complementary oligonucleotides with appropriate sticky end overhangs were annealed and ligated (T4 ligase NEB # M0202M) into the BsmBI- digested plasmid backbone.

    Article Title: Cell-free genomics reveals fundamental regulatory principles of the Mycobacterium tuberculosis transcription cycle.
    Article Snippet: Article Cell-free genomics reveals fundamental regulatory principles of the Mycobacterium tuberculosis transcription cycle

    Article Title: An activator of a two-component system controls cell separation and intrinsic drug resistance in Mycobacterium tuberculosis
    Article Snippet: Individual CRISPRi plasmids were cloned as described previously using Addgene plasmid 166886 [ ]. .. Briefly, the CRISPRi plasmid backbone was digested with BsmBI-v2 (NEB R0739L) and gel purified. sgRNAs were designed to target the non-template strand of the target gene ORF. .. For each individual sgRNA, two complementary oligonucleotides with appropriate sticky end overhangs were annealed and ligated (T4 ligase NEB M0202M) into the BsmBI-digested plasmid backbone.

    Article Title: Mapping unsolved lipidomes accelerates lipid discovery in major bacterial pathogens
    Article Snippet: Individual CRISPRi plasmids were generated following the protocol outlined in ( ) using Addgene plasmid #163631. .. Briefly, the CRISPRi plasmid backbone was digested with BsmBI-v2 (NEB #R0739L) and subsequently gel purified. sgRNAs were designed to target the non-template strand of the target gene ORF. .. For each sgRNA, two complementary oligonucleotides with compatible sticky-end overhangs were annealed and ligated into the BsmBI-digested plasmid backbone using T4 DNA ligase (NEB #M0202M).



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    CRISPRi-mediated silencing of the cwlM gene in M. smegmatis and resulting phenotypical and mRNA expression differences. a sgRNA-mediated repression of the cwlM gene in M. smegmatis , detailing target site (+ 467 bps) and the employed PAM strength (PAM14). b The local context of the cwlM gene in M. smegmatis . The cwlM gene is non-operonic, being preceded by the trx gene and succeeded by MSMEG_6936 , which is transcribed in the opposite direction. c Relative gene expression of cwlM normalized to sigA , with (stripped bars) and without (smooth bars) 100 ng/mL of ATc treatment for 6 h, evaluated through qRT-PCR assays ( n = 2). <t>PLJR962</t> is a non-targeting empty vector control. The mRNA expression levels of the calibrator sample ( M. smegmatis WT) are shown as dashed lines. The error bars represent the standard error of the mean (SEM). Comparisons between multiple groups were made using one-way ANOVA, with significance levels: * P < 0.05; ** P < 0.01. d Spotting dilution assays of the negative controls ( M. smegmatis WT, PLJR962) and of the cwlM knockdown mutant ( n = 3). The first spots were normalized to an OD 600 of 0.001, and the subsequent spots correspond to two-fold serial dilutions
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    Image Search Results


    CRISPRi-mediated silencing of the cwlM gene in M. smegmatis and resulting phenotypical and mRNA expression differences. a sgRNA-mediated repression of the cwlM gene in M. smegmatis , detailing target site (+ 467 bps) and the employed PAM strength (PAM14). b The local context of the cwlM gene in M. smegmatis . The cwlM gene is non-operonic, being preceded by the trx gene and succeeded by MSMEG_6936 , which is transcribed in the opposite direction. c Relative gene expression of cwlM normalized to sigA , with (stripped bars) and without (smooth bars) 100 ng/mL of ATc treatment for 6 h, evaluated through qRT-PCR assays ( n = 2). PLJR962 is a non-targeting empty vector control. The mRNA expression levels of the calibrator sample ( M. smegmatis WT) are shown as dashed lines. The error bars represent the standard error of the mean (SEM). Comparisons between multiple groups were made using one-way ANOVA, with significance levels: * P < 0.05; ** P < 0.01. d Spotting dilution assays of the negative controls ( M. smegmatis WT, PLJR962) and of the cwlM knockdown mutant ( n = 3). The first spots were normalized to an OD 600 of 0.001, and the subsequent spots correspond to two-fold serial dilutions

    Journal: BMC Microbiology

    Article Title: Effects of CwlM, a peptidoglycan synthesis regulator, on beta-lactam tolerance and host-pathogen interactions

    doi: 10.1186/s12866-025-04548-6

    Figure Lengend Snippet: CRISPRi-mediated silencing of the cwlM gene in M. smegmatis and resulting phenotypical and mRNA expression differences. a sgRNA-mediated repression of the cwlM gene in M. smegmatis , detailing target site (+ 467 bps) and the employed PAM strength (PAM14). b The local context of the cwlM gene in M. smegmatis . The cwlM gene is non-operonic, being preceded by the trx gene and succeeded by MSMEG_6936 , which is transcribed in the opposite direction. c Relative gene expression of cwlM normalized to sigA , with (stripped bars) and without (smooth bars) 100 ng/mL of ATc treatment for 6 h, evaluated through qRT-PCR assays ( n = 2). PLJR962 is a non-targeting empty vector control. The mRNA expression levels of the calibrator sample ( M. smegmatis WT) are shown as dashed lines. The error bars represent the standard error of the mean (SEM). Comparisons between multiple groups were made using one-way ANOVA, with significance levels: * P < 0.05; ** P < 0.01. d Spotting dilution assays of the negative controls ( M. smegmatis WT, PLJR962) and of the cwlM knockdown mutant ( n = 3). The first spots were normalized to an OD 600 of 0.001, and the subsequent spots correspond to two-fold serial dilutions

    Article Snippet: Escherichia coli ( E. coli ) strains were grown in Luria-Bertani (LB) media (Merck), overnight (ON) at 37 °C, to amplify the CRISPRi backbone PLJR962 (Addgene #115162) and for cloning.

    Techniques: Expressing, Gene Expression, Quantitative RT-PCR, Plasmid Preparation, Control, Knockdown, Mutagenesis

    Antibiotic susceptibility assays of the negative controls and the cwlM knockdown mutant. Heatmap of the fold differences in the minimum inhibitory concentrations (MICs; in µg/mL) between the cwlM knockdown mutant, the non-targeting control PLJR962 and M. smegmatis WT, with and without 100 ng/mL of ATc. The red shades depict an increase in the MIC while the blue shades depict a decrease in the MIC, when compared to the WT strain. AMX, amoxicillin; CLA, clavulanate; CTX, cefotaxime; EMB, ethambutol; INH, isoniazid; MEM, meropenem; VAN, vancomycin

    Journal: BMC Microbiology

    Article Title: Effects of CwlM, a peptidoglycan synthesis regulator, on beta-lactam tolerance and host-pathogen interactions

    doi: 10.1186/s12866-025-04548-6

    Figure Lengend Snippet: Antibiotic susceptibility assays of the negative controls and the cwlM knockdown mutant. Heatmap of the fold differences in the minimum inhibitory concentrations (MICs; in µg/mL) between the cwlM knockdown mutant, the non-targeting control PLJR962 and M. smegmatis WT, with and without 100 ng/mL of ATc. The red shades depict an increase in the MIC while the blue shades depict a decrease in the MIC, when compared to the WT strain. AMX, amoxicillin; CLA, clavulanate; CTX, cefotaxime; EMB, ethambutol; INH, isoniazid; MEM, meropenem; VAN, vancomycin

    Article Snippet: Escherichia coli ( E. coli ) strains were grown in Luria-Bertani (LB) media (Merck), overnight (ON) at 37 °C, to amplify the CRISPRi backbone PLJR962 (Addgene #115162) and for cloning.

    Techniques: Knockdown, Mutagenesis, Control