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spcas9 coding sequences  (Addgene inc)


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    Addgene inc spcas9 coding sequences
    Fig. 2 The modified 3TC scaffold boosts <t>SpCas9</t> gRNA expression levels compared to the original 4T scaffold. (A) DNA sequence of the 4T and modified 3TC scaffolds. (B) Relative quantification (RQ) of mDmd gRNA delivered by nucleofection of PX459.V2 (4T) to C2C12 cells, by qRT-PCR. Mean Log2RQ ± 95% CI; n = 3. One-way ANOVA with Dunnett’s multiple comparisons test was performed on ΔΔCT values, **p ≤ 0.01, ***p ≤ 0.001. (C, D, E) Comparison of the relative quantities of mDmd Sp gRNA, delivered by PX459.V2, pdg459.V2 (2 × 4T) and PX459.V3 (3TC), measured by qRT-PCR. Mean Log2RQ ± 95% CI; n = 3. One-way ANOVA with Tukey’s multiple comparisons test performed on ΔΔCT values, **p ≤ 0.01, ***p ≤ 0.001
    Spcas9 Coding Sequences, supplied by Addgene inc, used in various techniques. Bioz Stars score: 96/100, based on 369 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/cas9+coding+sequence/SP-Cas9+(Plasmid+%2362731)/pm39939860-134-6-17
    Average 96 stars, based on 369 article reviews
    spcas9 coding sequences - by Bioz Stars, 2026-10
    96/100 stars

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    1) Product Images from "Optimal SpCas9- and SaCas9-mediated gene editing by enhancing gRNA transcript levels through scaffold poly-T tract reduction."

    Article Title: Optimal SpCas9- and SaCas9-mediated gene editing by enhancing gRNA transcript levels through scaffold poly-T tract reduction.

    Journal: BMC genomics

    doi: 10.1186/s12864-025-11317-2

    Fig. 2 The modified 3TC scaffold boosts SpCas9 gRNA expression levels compared to the original 4T scaffold. (A) DNA sequence of the 4T and modified 3TC scaffolds. (B) Relative quantification (RQ) of mDmd gRNA delivered by nucleofection of PX459.V2 (4T) to C2C12 cells, by qRT-PCR. Mean Log2RQ ± 95% CI; n = 3. One-way ANOVA with Dunnett’s multiple comparisons test was performed on ΔΔCT values, **p ≤ 0.01, ***p ≤ 0.001. (C, D, E) Comparison of the relative quantities of mDmd Sp gRNA, delivered by PX459.V2, pdg459.V2 (2 × 4T) and PX459.V3 (3TC), measured by qRT-PCR. Mean Log2RQ ± 95% CI; n = 3. One-way ANOVA with Tukey’s multiple comparisons test performed on ΔΔCT values, **p ≤ 0.01, ***p ≤ 0.001
    Figure Legend Snippet: Fig. 2 The modified 3TC scaffold boosts SpCas9 gRNA expression levels compared to the original 4T scaffold. (A) DNA sequence of the 4T and modified 3TC scaffolds. (B) Relative quantification (RQ) of mDmd gRNA delivered by nucleofection of PX459.V2 (4T) to C2C12 cells, by qRT-PCR. Mean Log2RQ ± 95% CI; n = 3. One-way ANOVA with Dunnett’s multiple comparisons test was performed on ΔΔCT values, **p ≤ 0.01, ***p ≤ 0.001. (C, D, E) Comparison of the relative quantities of mDmd Sp gRNA, delivered by PX459.V2, pdg459.V2 (2 × 4T) and PX459.V3 (3TC), measured by qRT-PCR. Mean Log2RQ ± 95% CI; n = 3. One-way ANOVA with Tukey’s multiple comparisons test performed on ΔΔCT values, **p ≤ 0.01, ***p ≤ 0.001

    Techniques Used: Modification, Expressing, Sequencing, Quantitative Proteomics, Quantitative RT-PCR, Comparison

    Fig. 5 Editing efficiencies of high-fidelity SpCas9s with the 3TC scaffold. Comparison of PX459.V2 SpCas9-HF1 (4T), PX459.V3 SpCas9-HF1 (3TC), PX459.V2 eSpCas9(1.1) (4T) and PX459.V3 eSpCas9(1.1) (3TC) plasmids de livered by lipofection at a (A) high and (B) low plasmid dose without puro mycin selection in HEK239T cells, assessed by deep amplicon sequencing. Mean ± SEM; n = 3. Two-way ANOVA with Šídák’s multiple comparisons test; *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001. (C) Editing efficiencies of hDMD-B in the G19 gRNA configuration with WT and high-fidelity Sp-Cas9 plasmids delivered by nucleofection with puromycin selection in HEK293Ts. Two- way ANOVA with Šídák’s multiple comparisons test; *p ≤ 0.05, ***p ≤ 0.001
    Figure Legend Snippet: Fig. 5 Editing efficiencies of high-fidelity SpCas9s with the 3TC scaffold. Comparison of PX459.V2 SpCas9-HF1 (4T), PX459.V3 SpCas9-HF1 (3TC), PX459.V2 eSpCas9(1.1) (4T) and PX459.V3 eSpCas9(1.1) (3TC) plasmids de livered by lipofection at a (A) high and (B) low plasmid dose without puro mycin selection in HEK239T cells, assessed by deep amplicon sequencing. Mean ± SEM; n = 3. Two-way ANOVA with Šídák’s multiple comparisons test; *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001. (C) Editing efficiencies of hDMD-B in the G19 gRNA configuration with WT and high-fidelity Sp-Cas9 plasmids delivered by nucleofection with puromycin selection in HEK293Ts. Two- way ANOVA with Šídák’s multiple comparisons test; *p ≤ 0.05, ***p ≤ 0.001

    Techniques Used: Comparison, Plasmid Preparation, Selection, Amplification, Sequencing

    Related Articles

    Plasmid Preparation:

    Article Title: The Protein Tyrosine Phosphatase CD45 promotes PMN Transepithelial Migration, Antimicrobial Function and Colonic Mucosal Repair
    Article Snippet: .. The SpCas9 of lentiCRISPR V2 (Addgene plasmid # 52961) was modified to generate a plasmid containing the high fidelity eSpCas9 1.1 ( ) termed lentiCRISPR eSpCAS9. ..

    Article Title: mRNA-engineered CRISPR-Cas epigenetic editors enable durable and efficient gene silencing in vivo
    Article Snippet: .. The mammalian expression plasmid for the initial SpCas9-OFF-EE V1 was obtained from Addgene (Addgene, #167981). ..

    Article Title: The BRCA1-A complex restricts replication fork reversal-dependent DNA repair in ATM deficient cells
    Article Snippet: .. For SpCas9, stable cell lines expressing SpCas9 were first generated by lentiviral transduction with lenti-SpCas9 hygro (Addgene, plasmid #104995), and subsequent selection with hygromycin (500 μg/ml). sgRNA guides were cloned into either lentiGuide-Puro (Addgene, plasmid #52963) or lentiGuide-neo (U6-sgRNA-P2A-Neo, reconstructed from Addgene plasmid #52963) vector digested with BsmBI-v2 (New England Biolabs) following the protocol described previously . sgRNA plasmids (2.7 μg) were packaged into lentivirus particles by co-transfecting with packaging plasmids psPAX2 (2 μg, Addgene, plasmid #12260) and pMD2.G (1.3 μg, Addgene, plasmid #12259) into HEK293T cells grown in a 6-well plate using lipofectamine 2000 (Invitrogen). .. Lentivirus supernatant was collected 48 h post-transfection, filtered through a 0.45-μm filter (Millipore) and subsequently used to transduce the target cells using polybrene (8 μg/ml, Sigma-Aldrich) by spin infection at 2000 rpm for 90 min.

    Article Title: An upstream open reading frame represses translation of the neuronal potassium channel KCNQ2
    Article Snippet: The digested fragments were then ligated using T4 DNA ligase (Promega), creating plasmid psgRNA-EGFP. .. The plasmid encoding the ABE8e adenine base editor fused to an spCas9 nickase was obtained as a generous gift from David Liu (Addgene Plasmid #138489). ..

    Article Title: Compositions for use in treating autosomal dominant BEST1-related retinopathies
    Article Snippet: .. In addition, the px458 plasmid carrying the sgRNA 581G-17nt sequence (SEQ ID NO: 13) expresses both, the SpCas9 and the red fluorescent protein mcherry (Addgene #64324), while the px458 plasmid carrying sgRNA 749A-20nt (SEQ ID NO: 55) expresses the SpCas9 as well as the green fluorescent protein GFP (Addgene #48138). ..

    Modification:

    Article Title: The Protein Tyrosine Phosphatase CD45 promotes PMN Transepithelial Migration, Antimicrobial Function and Colonic Mucosal Repair
    Article Snippet: .. The SpCas9 of lentiCRISPR V2 (Addgene plasmid # 52961) was modified to generate a plasmid containing the high fidelity eSpCas9 1.1 ( ) termed lentiCRISPR eSpCAS9. ..

    Expressing:

    Article Title: AcrIIA7 hijacks tracrRNA to block CRISPR-Cas system.
    Article Snippet: .. Additionally, expression constructs for Streptococcus pyogenes Cas9 (#62731), Neisseria meningitides Cas9 (#71474), Haemophilus parainfluenzae Cas9 (#121540), and Simonsiella muelleri Cas9 (#121541) were procured from Addgene. ..

    Article Title: mRNA-engineered CRISPR-Cas epigenetic editors enable durable and efficient gene silencing in vivo
    Article Snippet: .. The mammalian expression plasmid for the initial SpCas9-OFF-EE V1 was obtained from Addgene (Addgene, #167981). ..

    Article Title: The BRCA1-A complex restricts replication fork reversal-dependent DNA repair in ATM deficient cells
    Article Snippet: .. For SpCas9, stable cell lines expressing SpCas9 were first generated by lentiviral transduction with lenti-SpCas9 hygro (Addgene, plasmid #104995), and subsequent selection with hygromycin (500 μg/ml). sgRNA guides were cloned into either lentiGuide-Puro (Addgene, plasmid #52963) or lentiGuide-neo (U6-sgRNA-P2A-Neo, reconstructed from Addgene plasmid #52963) vector digested with BsmBI-v2 (New England Biolabs) following the protocol described previously . sgRNA plasmids (2.7 μg) were packaged into lentivirus particles by co-transfecting with packaging plasmids psPAX2 (2 μg, Addgene, plasmid #12260) and pMD2.G (1.3 μg, Addgene, plasmid #12259) into HEK293T cells grown in a 6-well plate using lipofectamine 2000 (Invitrogen). .. Lentivirus supernatant was collected 48 h post-transfection, filtered through a 0.45-μm filter (Millipore) and subsequently used to transduce the target cells using polybrene (8 μg/ml, Sigma-Aldrich) by spin infection at 2000 rpm for 90 min.

    Construct:

    Article Title: AcrIIA7 hijacks tracrRNA to block CRISPR-Cas system.
    Article Snippet: .. Additionally, expression constructs for Streptococcus pyogenes Cas9 (#62731), Neisseria meningitides Cas9 (#71474), Haemophilus parainfluenzae Cas9 (#121540), and Simonsiella muelleri Cas9 (#121541) were procured from Addgene. ..

    Stable Transfection:

    Article Title: The BRCA1-A complex restricts replication fork reversal-dependent DNA repair in ATM deficient cells
    Article Snippet: .. For SpCas9, stable cell lines expressing SpCas9 were first generated by lentiviral transduction with lenti-SpCas9 hygro (Addgene, plasmid #104995), and subsequent selection with hygromycin (500 μg/ml). sgRNA guides were cloned into either lentiGuide-Puro (Addgene, plasmid #52963) or lentiGuide-neo (U6-sgRNA-P2A-Neo, reconstructed from Addgene plasmid #52963) vector digested with BsmBI-v2 (New England Biolabs) following the protocol described previously . sgRNA plasmids (2.7 μg) were packaged into lentivirus particles by co-transfecting with packaging plasmids psPAX2 (2 μg, Addgene, plasmid #12260) and pMD2.G (1.3 μg, Addgene, plasmid #12259) into HEK293T cells grown in a 6-well plate using lipofectamine 2000 (Invitrogen). .. Lentivirus supernatant was collected 48 h post-transfection, filtered through a 0.45-μm filter (Millipore) and subsequently used to transduce the target cells using polybrene (8 μg/ml, Sigma-Aldrich) by spin infection at 2000 rpm for 90 min.

    Generated:

    Article Title: The BRCA1-A complex restricts replication fork reversal-dependent DNA repair in ATM deficient cells
    Article Snippet: .. For SpCas9, stable cell lines expressing SpCas9 were first generated by lentiviral transduction with lenti-SpCas9 hygro (Addgene, plasmid #104995), and subsequent selection with hygromycin (500 μg/ml). sgRNA guides were cloned into either lentiGuide-Puro (Addgene, plasmid #52963) or lentiGuide-neo (U6-sgRNA-P2A-Neo, reconstructed from Addgene plasmid #52963) vector digested with BsmBI-v2 (New England Biolabs) following the protocol described previously . sgRNA plasmids (2.7 μg) were packaged into lentivirus particles by co-transfecting with packaging plasmids psPAX2 (2 μg, Addgene, plasmid #12260) and pMD2.G (1.3 μg, Addgene, plasmid #12259) into HEK293T cells grown in a 6-well plate using lipofectamine 2000 (Invitrogen). .. Lentivirus supernatant was collected 48 h post-transfection, filtered through a 0.45-μm filter (Millipore) and subsequently used to transduce the target cells using polybrene (8 μg/ml, Sigma-Aldrich) by spin infection at 2000 rpm for 90 min.

    Transduction:

    Article Title: The BRCA1-A complex restricts replication fork reversal-dependent DNA repair in ATM deficient cells
    Article Snippet: .. For SpCas9, stable cell lines expressing SpCas9 were first generated by lentiviral transduction with lenti-SpCas9 hygro (Addgene, plasmid #104995), and subsequent selection with hygromycin (500 μg/ml). sgRNA guides were cloned into either lentiGuide-Puro (Addgene, plasmid #52963) or lentiGuide-neo (U6-sgRNA-P2A-Neo, reconstructed from Addgene plasmid #52963) vector digested with BsmBI-v2 (New England Biolabs) following the protocol described previously . sgRNA plasmids (2.7 μg) were packaged into lentivirus particles by co-transfecting with packaging plasmids psPAX2 (2 μg, Addgene, plasmid #12260) and pMD2.G (1.3 μg, Addgene, plasmid #12259) into HEK293T cells grown in a 6-well plate using lipofectamine 2000 (Invitrogen). .. Lentivirus supernatant was collected 48 h post-transfection, filtered through a 0.45-μm filter (Millipore) and subsequently used to transduce the target cells using polybrene (8 μg/ml, Sigma-Aldrich) by spin infection at 2000 rpm for 90 min.

    Selection:

    Article Title: The BRCA1-A complex restricts replication fork reversal-dependent DNA repair in ATM deficient cells
    Article Snippet: .. For SpCas9, stable cell lines expressing SpCas9 were first generated by lentiviral transduction with lenti-SpCas9 hygro (Addgene, plasmid #104995), and subsequent selection with hygromycin (500 μg/ml). sgRNA guides were cloned into either lentiGuide-Puro (Addgene, plasmid #52963) or lentiGuide-neo (U6-sgRNA-P2A-Neo, reconstructed from Addgene plasmid #52963) vector digested with BsmBI-v2 (New England Biolabs) following the protocol described previously . sgRNA plasmids (2.7 μg) were packaged into lentivirus particles by co-transfecting with packaging plasmids psPAX2 (2 μg, Addgene, plasmid #12260) and pMD2.G (1.3 μg, Addgene, plasmid #12259) into HEK293T cells grown in a 6-well plate using lipofectamine 2000 (Invitrogen). .. Lentivirus supernatant was collected 48 h post-transfection, filtered through a 0.45-μm filter (Millipore) and subsequently used to transduce the target cells using polybrene (8 μg/ml, Sigma-Aldrich) by spin infection at 2000 rpm for 90 min.

    Clone Assay:

    Article Title: The BRCA1-A complex restricts replication fork reversal-dependent DNA repair in ATM deficient cells
    Article Snippet: .. For SpCas9, stable cell lines expressing SpCas9 were first generated by lentiviral transduction with lenti-SpCas9 hygro (Addgene, plasmid #104995), and subsequent selection with hygromycin (500 μg/ml). sgRNA guides were cloned into either lentiGuide-Puro (Addgene, plasmid #52963) or lentiGuide-neo (U6-sgRNA-P2A-Neo, reconstructed from Addgene plasmid #52963) vector digested with BsmBI-v2 (New England Biolabs) following the protocol described previously . sgRNA plasmids (2.7 μg) were packaged into lentivirus particles by co-transfecting with packaging plasmids psPAX2 (2 μg, Addgene, plasmid #12260) and pMD2.G (1.3 μg, Addgene, plasmid #12259) into HEK293T cells grown in a 6-well plate using lipofectamine 2000 (Invitrogen). .. Lentivirus supernatant was collected 48 h post-transfection, filtered through a 0.45-μm filter (Millipore) and subsequently used to transduce the target cells using polybrene (8 μg/ml, Sigma-Aldrich) by spin infection at 2000 rpm for 90 min.

    Sequencing:

    Article Title: Compositions for use in treating autosomal dominant BEST1-related retinopathies
    Article Snippet: .. In addition, the px458 plasmid carrying the sgRNA 581G-17nt sequence (SEQ ID NO: 13) expresses both, the SpCas9 and the red fluorescent protein mcherry (Addgene #64324), while the px458 plasmid carrying sgRNA 749A-20nt (SEQ ID NO: 55) expresses the SpCas9 as well as the green fluorescent protein GFP (Addgene #48138). ..

    Polymerase Chain Reaction:

    Article Title: In vitro and in vivo base editing of CCR5 in hematopoietic stem cells confers HIV-1 resistance.
    Article Snippet: This is a PDF of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability.. This version will undergo additional copyediting, typesetting and review before it is published in its final form.. As such, this version is no longer the Accepted Manuscript, but it is not yet the definitive Version of Record; we are providing this early version to give early visibility of the article.

    Ligation:

    Article Title: In vitro and in vivo base editing of CCR5 in hematopoietic stem cells confers HIV-1 resistance.
    Article Snippet: This is a PDF of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability.. This version will undergo additional copyediting, typesetting and review before it is published in its final form.. As such, this version is no longer the Accepted Manuscript, but it is not yet the definitive Version of Record; we are providing this early version to give early visibility of the article.



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    Image Search Results


    Fig. 2 The modified 3TC scaffold boosts SpCas9 gRNA expression levels compared to the original 4T scaffold. (A) DNA sequence of the 4T and modified 3TC scaffolds. (B) Relative quantification (RQ) of mDmd gRNA delivered by nucleofection of PX459.V2 (4T) to C2C12 cells, by qRT-PCR. Mean Log2RQ ± 95% CI; n = 3. One-way ANOVA with Dunnett’s multiple comparisons test was performed on ΔΔCT values, **p ≤ 0.01, ***p ≤ 0.001. (C, D, E) Comparison of the relative quantities of mDmd Sp gRNA, delivered by PX459.V2, pdg459.V2 (2 × 4T) and PX459.V3 (3TC), measured by qRT-PCR. Mean Log2RQ ± 95% CI; n = 3. One-way ANOVA with Tukey’s multiple comparisons test performed on ΔΔCT values, **p ≤ 0.01, ***p ≤ 0.001

    Journal: BMC genomics

    Article Title: Optimal SpCas9- and SaCas9-mediated gene editing by enhancing gRNA transcript levels through scaffold poly-T tract reduction.

    doi: 10.1186/s12864-025-11317-2

    Figure Lengend Snippet: Fig. 2 The modified 3TC scaffold boosts SpCas9 gRNA expression levels compared to the original 4T scaffold. (A) DNA sequence of the 4T and modified 3TC scaffolds. (B) Relative quantification (RQ) of mDmd gRNA delivered by nucleofection of PX459.V2 (4T) to C2C12 cells, by qRT-PCR. Mean Log2RQ ± 95% CI; n = 3. One-way ANOVA with Dunnett’s multiple comparisons test was performed on ΔΔCT values, **p ≤ 0.01, ***p ≤ 0.001. (C, D, E) Comparison of the relative quantities of mDmd Sp gRNA, delivered by PX459.V2, pdg459.V2 (2 × 4T) and PX459.V3 (3TC), measured by qRT-PCR. Mean Log2RQ ± 95% CI; n = 3. One-way ANOVA with Tukey’s multiple comparisons test performed on ΔΔCT values, **p ≤ 0.01, ***p ≤ 0.001

    Article Snippet: SaCas9Puro.V2 was generated by replacing the SpCas9 coding sequences of PX459.V2 with SaCas9 coding sequences from PX601 (Addgene #61591).

    Techniques: Modification, Expressing, Sequencing, Quantitative Proteomics, Quantitative RT-PCR, Comparison

    Fig. 5 Editing efficiencies of high-fidelity SpCas9s with the 3TC scaffold. Comparison of PX459.V2 SpCas9-HF1 (4T), PX459.V3 SpCas9-HF1 (3TC), PX459.V2 eSpCas9(1.1) (4T) and PX459.V3 eSpCas9(1.1) (3TC) plasmids de livered by lipofection at a (A) high and (B) low plasmid dose without puro mycin selection in HEK239T cells, assessed by deep amplicon sequencing. Mean ± SEM; n = 3. Two-way ANOVA with Šídák’s multiple comparisons test; *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001. (C) Editing efficiencies of hDMD-B in the G19 gRNA configuration with WT and high-fidelity Sp-Cas9 plasmids delivered by nucleofection with puromycin selection in HEK293Ts. Two- way ANOVA with Šídák’s multiple comparisons test; *p ≤ 0.05, ***p ≤ 0.001

    Journal: BMC genomics

    Article Title: Optimal SpCas9- and SaCas9-mediated gene editing by enhancing gRNA transcript levels through scaffold poly-T tract reduction.

    doi: 10.1186/s12864-025-11317-2

    Figure Lengend Snippet: Fig. 5 Editing efficiencies of high-fidelity SpCas9s with the 3TC scaffold. Comparison of PX459.V2 SpCas9-HF1 (4T), PX459.V3 SpCas9-HF1 (3TC), PX459.V2 eSpCas9(1.1) (4T) and PX459.V3 eSpCas9(1.1) (3TC) plasmids de livered by lipofection at a (A) high and (B) low plasmid dose without puro mycin selection in HEK239T cells, assessed by deep amplicon sequencing. Mean ± SEM; n = 3. Two-way ANOVA with Šídák’s multiple comparisons test; *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001. (C) Editing efficiencies of hDMD-B in the G19 gRNA configuration with WT and high-fidelity Sp-Cas9 plasmids delivered by nucleofection with puromycin selection in HEK293Ts. Two- way ANOVA with Šídák’s multiple comparisons test; *p ≤ 0.05, ***p ≤ 0.001

    Article Snippet: SaCas9Puro.V2 was generated by replacing the SpCas9 coding sequences of PX459.V2 with SaCas9 coding sequences from PX601 (Addgene #61591).

    Techniques: Comparison, Plasmid Preparation, Selection, Amplification, Sequencing