px601 aav cmv nls sacas9 nls 3xha bghpa u6 bsai sgrna plasmid 61591 Search Results


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    Addgene inc px601 plasmid
    Px601 Plasmid, supplied by Addgene inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc guide rna sequences targeting ecel1
    Guide Rna Sequences Targeting Ecel1, supplied by Addgene inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc u6 bsai sgrna
    In v i tro knock-down of human RHO- T17M expression. (A) Schematic view of construction of 293T stably expressing human RHO protein and transfection of <t>pX601-EFS-SaCas9-U6-sgRNA</t> (SaCas9/SgRNA) plasmid. (B) T7E1 assay indicated that SaCas9/17-Sg1 and SaCas9/17-Sg2 were appeared to cut the mutant sequence specifically, the full-length amplicon was 760bp, the two truncated amplicons were 510 bp and 250 bp, respectively. (C) The cutting efficacy of two sgRNAs with SaCas9 determined by TA & Sanger sequencing in 293T cells. (D) Rhodopsin expression reduction was determined by western blotting in 293T cells transfected with RHO -T17M-pEGFPN1(RHO17 cells) transfected with 17-Sg1 and Sg2 plasmid, comparing to the RHO wt cells with 17-Sg1 and Sg2 plasmid.
    U6 Bsai Sgrna, supplied by Addgene inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/u6 bsai sgrna/product/Addgene inc
    Average 86 stars, based on 1 article reviews
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    u6 bsai sgrna - by Bioz Stars, 2023-02
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    Addgene inc px601 aav cmv
    In v i tro knock-down of human RHO- T17M expression. (A) Schematic view of construction of 293T stably expressing human RHO protein and transfection of <t>pX601-EFS-SaCas9-U6-sgRNA</t> (SaCas9/SgRNA) plasmid. (B) T7E1 assay indicated that SaCas9/17-Sg1 and SaCas9/17-Sg2 were appeared to cut the mutant sequence specifically, the full-length amplicon was 760bp, the two truncated amplicons were 510 bp and 250 bp, respectively. (C) The cutting efficacy of two sgRNAs with SaCas9 determined by TA & Sanger sequencing in 293T cells. (D) Rhodopsin expression reduction was determined by western blotting in 293T cells transfected with RHO -T17M-pEGFPN1(RHO17 cells) transfected with 17-Sg1 and Sg2 plasmid, comparing to the RHO wt cells with 17-Sg1 and Sg2 plasmid.
    Px601 Aav Cmv, supplied by Addgene inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/px601 aav cmv/product/Addgene inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    px601 aav cmv - by Bioz Stars, 2023-02
    86/100 stars
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    Addgene inc px601
    Key Resources Table
    Px601, supplied by Addgene inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/px601/product/Addgene inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    px601 - by Bioz Stars, 2023-02
    86/100 stars
      Buy from Supplier

    Image Search Results


    In v i tro knock-down of human RHO- T17M expression. (A) Schematic view of construction of 293T stably expressing human RHO protein and transfection of pX601-EFS-SaCas9-U6-sgRNA (SaCas9/SgRNA) plasmid. (B) T7E1 assay indicated that SaCas9/17-Sg1 and SaCas9/17-Sg2 were appeared to cut the mutant sequence specifically, the full-length amplicon was 760bp, the two truncated amplicons were 510 bp and 250 bp, respectively. (C) The cutting efficacy of two sgRNAs with SaCas9 determined by TA & Sanger sequencing in 293T cells. (D) Rhodopsin expression reduction was determined by western blotting in 293T cells transfected with RHO -T17M-pEGFPN1(RHO17 cells) transfected with 17-Sg1 and Sg2 plasmid, comparing to the RHO wt cells with 17-Sg1 and Sg2 plasmid.

    Journal: bioRxiv

    Article Title: Allele-specific gene editing approach for vision loss restoration in RHO -associated Retinitis Pigmentosa

    doi: 10.1101/2022.11.16.516784

    Figure Lengend Snippet: In v i tro knock-down of human RHO- T17M expression. (A) Schematic view of construction of 293T stably expressing human RHO protein and transfection of pX601-EFS-SaCas9-U6-sgRNA (SaCas9/SgRNA) plasmid. (B) T7E1 assay indicated that SaCas9/17-Sg1 and SaCas9/17-Sg2 were appeared to cut the mutant sequence specifically, the full-length amplicon was 760bp, the two truncated amplicons were 510 bp and 250 bp, respectively. (C) The cutting efficacy of two sgRNAs with SaCas9 determined by TA & Sanger sequencing in 293T cells. (D) Rhodopsin expression reduction was determined by western blotting in 293T cells transfected with RHO -T17M-pEGFPN1(RHO17 cells) transfected with 17-Sg1 and Sg2 plasmid, comparing to the RHO wt cells with 17-Sg1 and Sg2 plasmid.

    Article Snippet: The selected sgRNA was cloned into the plasmid pX601-AAVCMV: NLS-SaCas9-NLS-3xHA-bGHpA; U6: BsaI-sgRNA (pX601, #61591, Addgene).

    Techniques: Expressing, Stable Transfection, Transfection, Plasmid Preparation, Mutagenesis, Sequencing, Amplification, Western Blot

    Determination of specificity and cutting efficiency of SaCas9 and sgRNA in induced pluripotent stem cells (iPSCs) from patients’ urine cells (UCs). (A)Photograph of UCs. Scale bar=100 μm. (B) Photograph of iPSCs. Scale bar = 200 μm. (C-D) Sanger sequencing results showed that P1 iPS had the heterozygous c.50C>T mutation. IF staining showed the iPSCs expressed human embryonic stem cell-specific surface antigens including NANOG (E), OCT4 (F), TRA-1-60 (G) and SSEA-4 (H) protein. (I) Chromosomal content analysis revealed a normal 46, XY karyotype of patient P1. (J) Schematic view of AAV-EFS-SaCas9-U6-sgRNA-p2a-Puro plasmid. (K) T7E1 assay indicated that SaCas9/17-Sg2 was appeared to cut the mutant sequence specifically. (L) The cutting efficacy of SaCas9/17-Sg2 is determined by TA & Sanger sequencing in iPSCs.

    Journal: bioRxiv

    Article Title: Allele-specific gene editing approach for vision loss restoration in RHO -associated Retinitis Pigmentosa

    doi: 10.1101/2022.11.16.516784

    Figure Lengend Snippet: Determination of specificity and cutting efficiency of SaCas9 and sgRNA in induced pluripotent stem cells (iPSCs) from patients’ urine cells (UCs). (A)Photograph of UCs. Scale bar=100 μm. (B) Photograph of iPSCs. Scale bar = 200 μm. (C-D) Sanger sequencing results showed that P1 iPS had the heterozygous c.50C>T mutation. IF staining showed the iPSCs expressed human embryonic stem cell-specific surface antigens including NANOG (E), OCT4 (F), TRA-1-60 (G) and SSEA-4 (H) protein. (I) Chromosomal content analysis revealed a normal 46, XY karyotype of patient P1. (J) Schematic view of AAV-EFS-SaCas9-U6-sgRNA-p2a-Puro plasmid. (K) T7E1 assay indicated that SaCas9/17-Sg2 was appeared to cut the mutant sequence specifically. (L) The cutting efficacy of SaCas9/17-Sg2 is determined by TA & Sanger sequencing in iPSCs.

    Article Snippet: The selected sgRNA was cloned into the plasmid pX601-AAVCMV: NLS-SaCas9-NLS-3xHA-bGHpA; U6: BsaI-sgRNA (pX601, #61591, Addgene).

    Techniques: Sequencing, Mutagenesis, Staining, Plasmid Preparation

    (A) (Top) AAV2/8 vectors expressing EGFP under the control of EFS promoter; (Bottom) AAV2/8 vectors expressing SaCas9 under the control of EFS promoter and sgRNA under the control of U6 promoter were schematized. (B) Fluorescein shows AAV vectors distribution following subretinal injection immediately. (C) Infected areas treated with AAV2/8-EFS-EGFP vector at dose of 1×10 9 vg per eye (1×10 9 dose) were labelled by GFP expression (GFP+ area). ONL = outer nuclear layer; OS = outer segment; IS = inner segment. Scale bar = 50 μm. (D) IF staining indicates GFP expresses in mouse retinas at 1m and 11 mpi treated with AAV2/8-EFS-EGFP vector at 3×10 9 dose. (E) The quantitative PCR analysis indicated the expression of SaCas9, 17-Sg2 and GFP in Rho wt/hum retinas at 3 mpi treated with AAV2/8-EFS-SaCas9-U6-17-Sg2 and AAV2/8-EFS-EGFP vector (1:1 mixture) at different doses. (F) Determination of SaCas9 expression by western blotting at 3 mpi at 1×10 9 and 3×10 9 dose. (Left) Mouse #h, i and j ( Rho wt/hum ) were injected with AAV2/8-EFS-SaCas9-U6-17-Sg2 and AAV2/8-EFS-EGFP vector (1:1 mixture) at 1×10 9 dose; (Right) Mouse #k, l, m and n ( Rho wt/hum ) were injected with AAV2/8-EFS-SaCas9-U6-17-Sg2 and AAV2/8-EFS-EGFP vector (1:1 mixture) at 3×10 9 dose.

    Journal: bioRxiv

    Article Title: Allele-specific gene editing approach for vision loss restoration in RHO -associated Retinitis Pigmentosa

    doi: 10.1101/2022.11.16.516784

    Figure Lengend Snippet: (A) (Top) AAV2/8 vectors expressing EGFP under the control of EFS promoter; (Bottom) AAV2/8 vectors expressing SaCas9 under the control of EFS promoter and sgRNA under the control of U6 promoter were schematized. (B) Fluorescein shows AAV vectors distribution following subretinal injection immediately. (C) Infected areas treated with AAV2/8-EFS-EGFP vector at dose of 1×10 9 vg per eye (1×10 9 dose) were labelled by GFP expression (GFP+ area). ONL = outer nuclear layer; OS = outer segment; IS = inner segment. Scale bar = 50 μm. (D) IF staining indicates GFP expresses in mouse retinas at 1m and 11 mpi treated with AAV2/8-EFS-EGFP vector at 3×10 9 dose. (E) The quantitative PCR analysis indicated the expression of SaCas9, 17-Sg2 and GFP in Rho wt/hum retinas at 3 mpi treated with AAV2/8-EFS-SaCas9-U6-17-Sg2 and AAV2/8-EFS-EGFP vector (1:1 mixture) at different doses. (F) Determination of SaCas9 expression by western blotting at 3 mpi at 1×10 9 and 3×10 9 dose. (Left) Mouse #h, i and j ( Rho wt/hum ) were injected with AAV2/8-EFS-SaCas9-U6-17-Sg2 and AAV2/8-EFS-EGFP vector (1:1 mixture) at 1×10 9 dose; (Right) Mouse #k, l, m and n ( Rho wt/hum ) were injected with AAV2/8-EFS-SaCas9-U6-17-Sg2 and AAV2/8-EFS-EGFP vector (1:1 mixture) at 3×10 9 dose.

    Article Snippet: The selected sgRNA was cloned into the plasmid pX601-AAVCMV: NLS-SaCas9-NLS-3xHA-bGHpA; U6: BsaI-sgRNA (pX601, #61591, Addgene).

    Techniques: Expressing, Injection, Infection, Plasmid Preparation, Staining, Real-time Polymerase Chain Reaction, Western Blot

    Identification of single-nucleotide variants (SNVs, A) and indels (C) in 293T cells transfected with pX601-EFS-SaCas9-U6-17-Sg2 (17-Sg2) plasmid at the WGS level. The type of SNVs (B) and indels (D) in 293T cells transfected with 17-Sg2 plasmid and untreated cells at the WGS level.

    Journal: bioRxiv

    Article Title: Allele-specific gene editing approach for vision loss restoration in RHO -associated Retinitis Pigmentosa

    doi: 10.1101/2022.11.16.516784

    Figure Lengend Snippet: Identification of single-nucleotide variants (SNVs, A) and indels (C) in 293T cells transfected with pX601-EFS-SaCas9-U6-17-Sg2 (17-Sg2) plasmid at the WGS level. The type of SNVs (B) and indels (D) in 293T cells transfected with 17-Sg2 plasmid and untreated cells at the WGS level.

    Article Snippet: The selected sgRNA was cloned into the plasmid pX601-AAVCMV: NLS-SaCas9-NLS-3xHA-bGHpA; U6: BsaI-sgRNA (pX601, #61591, Addgene).

    Techniques: Transfection, Plasmid Preparation

    Key Resources Table

    Journal: Cell reports

    Article Title: CRISPR-READI: Efficient generation of knock-in mice by CRISPR RNP Electroporation and AAV Donor Infection

    doi: 10.1016/j.celrep.2019.05.103

    Figure Lengend Snippet: Key Resources Table

    Article Snippet: A plasmid derivative of pX601 (Addgene #61591) containing a nanoluciferase cassette was digested with HindIII and ligated to the pGEM stuffer sequence to generate pX601-long.

    Techniques: Recombinant, Software