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sacas9 site 2 sgrna  (Addgene inc)


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    Addgene inc sacas9 site 2 sgrna
    Sacas9 Site 2 Sgrna, supplied by Addgene inc, used in various techniques. Bioz Stars score: 96/100, based on 164 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/pspgrna/pSPgRNA+(Plasmid+%2347108)/pm41832601-359-15-34
    Average 96 stars, based on 164 article reviews
    sacas9 site 2 sgrna - by Bioz Stars, 2026-10
    96/100 stars

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    Related Articles

    Single Cell:

    Article Title: Highly efficient XIST reactivation in female hPSC by transient dual inhibition of TP53 and DNA methylation during Cas9 mediated genome editing
    Article Snippet: hPSCs were treated with GSK3685032 (#HY-139664; MedChemExpress, NJ, USA) for indicated periods and concentrations in each figure. .. These hPSCs were dissociated into single cell using TrypLE Express (#12605010; Thermo Fisher Scientific) and subjected to transfection (Neon Transfection System, Thermo Fisher Scientific) of Cas9-T2A-EGFP-ires-puro (a gift from Timo Otonkoski, #78311; Addgene), pCE-mp53DD (a gift from Shinya Yamanaka Plasmid #41856), pSPgRNA (a gift from Charles Gersbach, #47108; Addgene), targeting XIST promoter (REF). after transfection, hPSCs were cultured in the presence of GSK3685032 or not for two days followed by FACS sorting (BD, Franklin Lakes, NJ, USA). ..

    Article Title: Highly efficient XIST reactivation in female hPSC by transient dual inhibition of TP53 and DNA methylation during Cas9 mediated genome editing
    Article Snippet: hPSCs were treated with GSK3685032 (#HY-139664; MedChemExpress, NJ, USA) for indicated periods and concentrations in each figure. .. These hPSCs were dissociated into single cell using TrypLE Express (#12605010; Thermo Fisher Scientific) and subjected to transfection (Neon Transfection System, Thermo Fisher Scientific) of Cas9-T2A-EGFP-ires-puro (a gift from Timo Otonkoski, #78311; Addgene), pCE-mp53DD (a gift from Shinya Yamanaka Plasmid #41856), pSPgRNA (a gift from Charles Gersbach, #47108; Addgene), targeting XIST promoter [ ]. .. After transfection, hPSCs were cultured in the presence of GSK3685032 or not for two days followed by FACS sorting (BD, Franklin Lakes , NJ, USA).

    Transfection:

    Article Title: Highly efficient XIST reactivation in female hPSC by transient dual inhibition of TP53 and DNA methylation during Cas9 mediated genome editing
    Article Snippet: hPSCs were treated with GSK3685032 (#HY-139664; MedChemExpress, NJ, USA) for indicated periods and concentrations in each figure. .. These hPSCs were dissociated into single cell using TrypLE Express (#12605010; Thermo Fisher Scientific) and subjected to transfection (Neon Transfection System, Thermo Fisher Scientific) of Cas9-T2A-EGFP-ires-puro (a gift from Timo Otonkoski, #78311; Addgene), pCE-mp53DD (a gift from Shinya Yamanaka Plasmid #41856), pSPgRNA (a gift from Charles Gersbach, #47108; Addgene), targeting XIST promoter (REF). after transfection, hPSCs were cultured in the presence of GSK3685032 or not for two days followed by FACS sorting (BD, Franklin Lakes, NJ, USA). ..

    Article Title: Highly efficient XIST reactivation in female hPSC by transient dual inhibition of TP53 and DNA methylation during Cas9 mediated genome editing
    Article Snippet: hPSCs were treated with GSK3685032 (#HY-139664; MedChemExpress, NJ, USA) for indicated periods and concentrations in each figure. .. These hPSCs were dissociated into single cell using TrypLE Express (#12605010; Thermo Fisher Scientific) and subjected to transfection (Neon Transfection System, Thermo Fisher Scientific) of Cas9-T2A-EGFP-ires-puro (a gift from Timo Otonkoski, #78311; Addgene), pCE-mp53DD (a gift from Shinya Yamanaka Plasmid #41856), pSPgRNA (a gift from Charles Gersbach, #47108; Addgene), targeting XIST promoter [ ]. .. After transfection, hPSCs were cultured in the presence of GSK3685032 or not for two days followed by FACS sorting (BD, Franklin Lakes , NJ, USA).

    Plasmid Preparation:

    Article Title: Highly efficient XIST reactivation in female hPSC by transient dual inhibition of TP53 and DNA methylation during Cas9 mediated genome editing
    Article Snippet: hPSCs were treated with GSK3685032 (#HY-139664; MedChemExpress, NJ, USA) for indicated periods and concentrations in each figure. .. These hPSCs were dissociated into single cell using TrypLE Express (#12605010; Thermo Fisher Scientific) and subjected to transfection (Neon Transfection System, Thermo Fisher Scientific) of Cas9-T2A-EGFP-ires-puro (a gift from Timo Otonkoski, #78311; Addgene), pCE-mp53DD (a gift from Shinya Yamanaka Plasmid #41856), pSPgRNA (a gift from Charles Gersbach, #47108; Addgene), targeting XIST promoter (REF). after transfection, hPSCs were cultured in the presence of GSK3685032 or not for two days followed by FACS sorting (BD, Franklin Lakes, NJ, USA). ..

    Article Title: Regulation of the cell surface expression of classical and non-classical MHC proteins by the human cytomegalovirus UL40 and rhesus cytomegalovirus Rh67 proteins
    Article Snippet: Transfections of HEK 293T cells were carried out in six-well plates, with cells at 50%–70% confluency, using GeneJuice (Merck) with 1 μg of total plasmid DNA per well, as per the manufacturer’s instructions. .. Guide RNAs for TAP1, TAP2, and HM13 [Table S2 ( )] were inserted into pspgRNA [ , Addgene plasmid #47108, a gift from Charles Gersbach]. .. HEK 293T cells were co-transfected with equal amounts of the pspgRNA plasmids and pCas9_GFP [ , Addgene plasmid #44719, a gift from Kiran Musunuru].

    Article Title: A microglia clonal inflammatory disorder in Alzheimer’s Disease
    Article Snippet: .. The sgRNA target sequence was cloned into the pSPgRNA (Addgene plasmid # 47108) Perez- Pinera et al., 2013 to make the gene targeting construct. .. To introduce the CBL C404Y variants, the WT hiPSC (C12) were dissociated using Accutase (Innovative Cell Technologies) and electroporated (1×106 cells per reaction) with 4 μg sgRNA- construct plasmid and 4 μg CBE enzyme coding vector BE3- FNLS (Addgene plasmid # 112671) (Zafra et al., 2018) using Lonza 4D- Nucleofector and the Nucleofector solution (Lonza V4XP- 3034) following our previously reported protocol (Zhong et al., 2020).

    Article Title: Spatiotemporal proteomics reveals dynamic antagonistic gradients shaping signalling waves
    Article Snippet: .. In agreement with the CRISPaint editing system we prepared 3 vectors: pSPgRNA (Addgene plasmid #47108) containing the RSPO3 sgRNA sequence (5’- AGTCAGCACTGTACACTAGA-3’); donor vector containing reporter cassette with the T2A-Achilles-NLS-PEST sequence and a selection cassette with Neomycin; and the corresponding Frame Selector pCAS9-mCherry-Frame +2. ..

    Article Title: Highly efficient XIST reactivation in female hPSC by transient dual inhibition of TP53 and DNA methylation during Cas9 mediated genome editing
    Article Snippet: hPSCs were treated with GSK3685032 (#HY-139664; MedChemExpress, NJ, USA) for indicated periods and concentrations in each figure. .. These hPSCs were dissociated into single cell using TrypLE Express (#12605010; Thermo Fisher Scientific) and subjected to transfection (Neon Transfection System, Thermo Fisher Scientific) of Cas9-T2A-EGFP-ires-puro (a gift from Timo Otonkoski, #78311; Addgene), pCE-mp53DD (a gift from Shinya Yamanaka Plasmid #41856), pSPgRNA (a gift from Charles Gersbach, #47108; Addgene), targeting XIST promoter [ ]. .. After transfection, hPSCs were cultured in the presence of GSK3685032 or not for two days followed by FACS sorting (BD, Franklin Lakes , NJ, USA).

    Cell Culture:

    Article Title: Highly efficient XIST reactivation in female hPSC by transient dual inhibition of TP53 and DNA methylation during Cas9 mediated genome editing
    Article Snippet: hPSCs were treated with GSK3685032 (#HY-139664; MedChemExpress, NJ, USA) for indicated periods and concentrations in each figure. .. These hPSCs were dissociated into single cell using TrypLE Express (#12605010; Thermo Fisher Scientific) and subjected to transfection (Neon Transfection System, Thermo Fisher Scientific) of Cas9-T2A-EGFP-ires-puro (a gift from Timo Otonkoski, #78311; Addgene), pCE-mp53DD (a gift from Shinya Yamanaka Plasmid #41856), pSPgRNA (a gift from Charles Gersbach, #47108; Addgene), targeting XIST promoter (REF). after transfection, hPSCs were cultured in the presence of GSK3685032 or not for two days followed by FACS sorting (BD, Franklin Lakes, NJ, USA). ..

    FACS:

    Article Title: Highly efficient XIST reactivation in female hPSC by transient dual inhibition of TP53 and DNA methylation during Cas9 mediated genome editing
    Article Snippet: hPSCs were treated with GSK3685032 (#HY-139664; MedChemExpress, NJ, USA) for indicated periods and concentrations in each figure. .. These hPSCs were dissociated into single cell using TrypLE Express (#12605010; Thermo Fisher Scientific) and subjected to transfection (Neon Transfection System, Thermo Fisher Scientific) of Cas9-T2A-EGFP-ires-puro (a gift from Timo Otonkoski, #78311; Addgene), pCE-mp53DD (a gift from Shinya Yamanaka Plasmid #41856), pSPgRNA (a gift from Charles Gersbach, #47108; Addgene), targeting XIST promoter (REF). after transfection, hPSCs were cultured in the presence of GSK3685032 or not for two days followed by FACS sorting (BD, Franklin Lakes, NJ, USA). ..

    Sequencing:

    Article Title: A microglia clonal inflammatory disorder in Alzheimer’s Disease
    Article Snippet: .. The sgRNA target sequence was cloned into the pSPgRNA (Addgene plasmid # 47108) Perez- Pinera et al., 2013 to make the gene targeting construct. .. To introduce the CBL C404Y variants, the WT hiPSC (C12) were dissociated using Accutase (Innovative Cell Technologies) and electroporated (1×106 cells per reaction) with 4 μg sgRNA- construct plasmid and 4 μg CBE enzyme coding vector BE3- FNLS (Addgene plasmid # 112671) (Zafra et al., 2018) using Lonza 4D- Nucleofector and the Nucleofector solution (Lonza V4XP- 3034) following our previously reported protocol (Zhong et al., 2020).

    Article Title: In vitro and in vivo expansion of CD33/HBG promoter-edited HSPCs with Mylotarg
    Article Snippet: GO (Mylotarg, Pfizer) was used for the in vivo selection of CD33-edited cells. .. The CD33 guide sequence 5′-CCCCACAGGGGCCCTGGCTA-3′ was inserted into the Bbs I site of pSPgRNA (Addgene, no. 47108), generating pSP-sgCD33. ..

    Article Title: Spatiotemporal proteomics reveals dynamic antagonistic gradients shaping signalling waves
    Article Snippet: .. In agreement with the CRISPaint editing system we prepared 3 vectors: pSPgRNA (Addgene plasmid #47108) containing the RSPO3 sgRNA sequence (5’- AGTCAGCACTGTACACTAGA-3’); donor vector containing reporter cassette with the T2A-Achilles-NLS-PEST sequence and a selection cassette with Neomycin; and the corresponding Frame Selector pCAS9-mCherry-Frame +2. ..

    Article Title: In vitro and in vivo expansion of CD33/HBG promoter-edited HSPCs with Mylotarg
    Article Snippet: GO (Mylotarg, Pfizer) was used for the in vivo selection of CD33-edited cells. .. Construction of HDAd-ABE8e-sgHBG#2-sgCD33DE2 The CD33 guide sequence 50-CCCCACAGGGGCCCTGGCTA-30 was inserted into the BbsI site of pSPgRNA (Addgene, no. 47108), generating pSP-sgCD33. ..

    Clone Assay:

    Article Title: A microglia clonal inflammatory disorder in Alzheimer’s Disease
    Article Snippet: .. The sgRNA target sequence was cloned into the pSPgRNA (Addgene plasmid # 47108) Perez- Pinera et al., 2013 to make the gene targeting construct. .. To introduce the CBL C404Y variants, the WT hiPSC (C12) were dissociated using Accutase (Innovative Cell Technologies) and electroporated (1×106 cells per reaction) with 4 μg sgRNA- construct plasmid and 4 μg CBE enzyme coding vector BE3- FNLS (Addgene plasmid # 112671) (Zafra et al., 2018) using Lonza 4D- Nucleofector and the Nucleofector solution (Lonza V4XP- 3034) following our previously reported protocol (Zhong et al., 2020).

    Construct:

    Article Title: A microglia clonal inflammatory disorder in Alzheimer’s Disease
    Article Snippet: .. The sgRNA target sequence was cloned into the pSPgRNA (Addgene plasmid # 47108) Perez- Pinera et al., 2013 to make the gene targeting construct. .. To introduce the CBL C404Y variants, the WT hiPSC (C12) were dissociated using Accutase (Innovative Cell Technologies) and electroporated (1×106 cells per reaction) with 4 μg sgRNA- construct plasmid and 4 μg CBE enzyme coding vector BE3- FNLS (Addgene plasmid # 112671) (Zafra et al., 2018) using Lonza 4D- Nucleofector and the Nucleofector solution (Lonza V4XP- 3034) following our previously reported protocol (Zhong et al., 2020).

    Polymerase Chain Reaction:

    Article Title: CRISPR-Cas-based activation of PPARGC1A boosts endogenous mitochondria and enhances cardiac function after myocardial infarction.
    Article Snippet: .. SadCas9-VPR, the hU6 promoter, and P2A-eGFP were polymerase chain reaction (PCR) amplified from SadCas9 VPR (Addgene, catalog no. 373494), pSPgRNA (Addgene, catalog no. 47108), and pCMV-T7-SpRY-P2A-EGFP (RTW4830; Addgene, catalog no. 139989), respectively. ..

    Amplification:

    Article Title: CRISPR-Cas-based activation of PPARGC1A boosts endogenous mitochondria and enhances cardiac function after myocardial infarction.
    Article Snippet: .. SadCas9-VPR, the hU6 promoter, and P2A-eGFP were polymerase chain reaction (PCR) amplified from SadCas9 VPR (Addgene, catalog no. 373494), pSPgRNA (Addgene, catalog no. 47108), and pCMV-T7-SpRY-P2A-EGFP (RTW4830; Addgene, catalog no. 139989), respectively. ..

    Selection:

    Article Title: Spatiotemporal proteomics reveals dynamic antagonistic gradients shaping signalling waves
    Article Snippet: .. In agreement with the CRISPaint editing system we prepared 3 vectors: pSPgRNA (Addgene plasmid #47108) containing the RSPO3 sgRNA sequence (5’- AGTCAGCACTGTACACTAGA-3’); donor vector containing reporter cassette with the T2A-Achilles-NLS-PEST sequence and a selection cassette with Neomycin; and the corresponding Frame Selector pCAS9-mCherry-Frame +2. ..



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    Image Search Results


    (A) Schematic of the PURPL IR reporter used in the screen to identify IR regulators. The pLCHKO vector contains the puromycin resistance gene used for selection. The transgene also contains the genomic sequences of exon 2 (161 bp), intron 2 (1,943 bp), and exon 3 (163 bp) of the PURPL gene under a Doxycycline-inducible promoter. A hybrid guide (hg)RNA for guiding SpCas9 and LbCas12a is expressed under a U6 promoter. A construct with a hgRNA targeting an intergenic region was used as a negative control. Top arrow: The hgRNA is cleaved by Cas12a to produce 2 guide RNAs for dual targeting of each protein coding gene, paralog targeting, and genetic interaction mapping. Bottom arrow: Upon intron splicing or intron retention, two different reads are generated. (B) Two different cell lines, stably expressing Cas9 and Cas12a, are used: HAP1, and RPE1. The pipeline before RNA-seq is depicted. (C) Heatmap showing single genes only (after removing gene pairs), whether identified individually or as part of a gene pair. The hits are ordered by the average ΔPIR values across HAP1 and RPE1, from lowest to highest. (D) Left: Gene ontology enrichment analysis of biological processes regulated by the hits of the screen. The most significant pathways involve mRNA processing and RNA splicing. Right: Gene ontology enrichment analysis of cellular components regulated by the hits of the screen. These components include mainly Spliceosomal Complex, small nuclear ribonucleoprotein complex and the Catalytic step 2 Spliceosome. (E) Venn diagram showing that there are 5 common hits which cause a reduction in the percentage of intron retention (ΔPIR) of the splicing reporter in the two cell lines. (F) Table showing the top 5 hits of the screen with a ΔPIR for each hit in each cell line and their full names.

    Journal: bioRxiv

    Article Title: Non-canonical function of the splicing activator U2AF2 in promoting intron retention in the lncRNAs PURPL and MALAT1

    doi: 10.64898/2026.02.19.706780

    Figure Lengend Snippet: (A) Schematic of the PURPL IR reporter used in the screen to identify IR regulators. The pLCHKO vector contains the puromycin resistance gene used for selection. The transgene also contains the genomic sequences of exon 2 (161 bp), intron 2 (1,943 bp), and exon 3 (163 bp) of the PURPL gene under a Doxycycline-inducible promoter. A hybrid guide (hg)RNA for guiding SpCas9 and LbCas12a is expressed under a U6 promoter. A construct with a hgRNA targeting an intergenic region was used as a negative control. Top arrow: The hgRNA is cleaved by Cas12a to produce 2 guide RNAs for dual targeting of each protein coding gene, paralog targeting, and genetic interaction mapping. Bottom arrow: Upon intron splicing or intron retention, two different reads are generated. (B) Two different cell lines, stably expressing Cas9 and Cas12a, are used: HAP1, and RPE1. The pipeline before RNA-seq is depicted. (C) Heatmap showing single genes only (after removing gene pairs), whether identified individually or as part of a gene pair. The hits are ordered by the average ΔPIR values across HAP1 and RPE1, from lowest to highest. (D) Left: Gene ontology enrichment analysis of biological processes regulated by the hits of the screen. The most significant pathways involve mRNA processing and RNA splicing. Right: Gene ontology enrichment analysis of cellular components regulated by the hits of the screen. These components include mainly Spliceosomal Complex, small nuclear ribonucleoprotein complex and the Catalytic step 2 Spliceosome. (E) Venn diagram showing that there are 5 common hits which cause a reduction in the percentage of intron retention (ΔPIR) of the splicing reporter in the two cell lines. (F) Table showing the top 5 hits of the screen with a ΔPIR for each hit in each cell line and their full names.

    Article Snippet: For HCT116 MALAT1 knockout, we used integration by non-homologous end joining, which was accomplished by introducing a simultaneous double-strand break in genomic DNA and in the targeting vectors (pCMV-Puro and pCMV-Hygro)., Plasmids encoding spCas9 and sgRNAs were obtained from Addgene (Plasmids #41815 and #47108).

    Techniques: Plasmid Preparation, Selection, Genomic Sequencing, Construct, Negative Control, Generated, Stable Transfection, Expressing, RNA Sequencing