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aav1 flexpkialpha ires nls mruby2  (Addgene inc)


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    Addgene inc aav1 flexpkialpha ires nls mruby2
    Aav1 Flexpkialpha Ires Nls Mruby2, supplied by Addgene inc, used in various techniques. Bioz Stars score: 91/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/mruby2/AAV-FLEX-PKIalpha-IRES-nls-mRuby2+(Plasmid+%2363059)/pm41906363-70-0-8
    Average 91 stars, based on 5 article reviews
    aav1 flexpkialpha ires nls mruby2 - by Bioz Stars, 2026-09
    91/100 stars

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    Related Articles

    Construct:

    Article Title: Calcineurin governs baseline and homeostatic regulations of non–rapid eye movement sleep in mice
    Article Snippet: Mouse open reading frames (ORFs) of CnAα (MR227082), CnAβ (MR208424), CnAγ (MR224578), and PKAα (MR205322) were purchased from OriGene Technologies. pAAV-Tre-3HA-CnAα/β/γ CA were constructed from pAAV-Tre-EGFP by replacing EGFP with ORF sequences of CnAα CA (AA 1-398), CnAβ CA (AA 1-408), and CnAγ CA (AA 1-391) mutants lacking the C-terminal autoinhibitory domain. pAAV-hSyn-3HA-RIαB was constructed by subcloning RIα G325D into pAAV-hSyn-EGFP (Addgene, 105539). pAAV-hSyn-DIO-3HA-RIαB was constructed by cloning RIα G325D into pAAV-hSyn-DIO-hM3D(Gq)-mCherry (Addgene, 44361). .. Addgene supplied pAAV-CaMKII-Cre (182736), and pAAV-Ple94-Cre (49125). pAAV-Dlx-Cre was constructed from pAAV-mDlx-NLS-mRuby2 (Addgene, 99130) by replacing mRuby2 with Cre. pCS2-3HA-SIK3 LxVPm2 was constructed by mutating two “LxVP” motifs to “AAAA” by site-directed mutagenesis. ..

    Article Title: PP2Acα regulates sleep amount and sleep homeostasis in mice
    Article Snippet: .. PP2Acα L309A was constructed by site-directed mutagenesis using specific primers that contain L309A mutations. pAAV-CaMKII-Cre (182736, Addgene) was purchased from Addgene. pAAV-mDlx-Cre was constructed by replacing mRuby2 with Cre from pAAV-mDlx-NLS-mRuby2 (99130, Addgene). ..

    Mutagenesis:

    Article Title: Calcineurin governs baseline and homeostatic regulations of non–rapid eye movement sleep in mice
    Article Snippet: Mouse open reading frames (ORFs) of CnAα (MR227082), CnAβ (MR208424), CnAγ (MR224578), and PKAα (MR205322) were purchased from OriGene Technologies. pAAV-Tre-3HA-CnAα/β/γ CA were constructed from pAAV-Tre-EGFP by replacing EGFP with ORF sequences of CnAα CA (AA 1-398), CnAβ CA (AA 1-408), and CnAγ CA (AA 1-391) mutants lacking the C-terminal autoinhibitory domain. pAAV-hSyn-3HA-RIαB was constructed by subcloning RIα G325D into pAAV-hSyn-EGFP (Addgene, 105539). pAAV-hSyn-DIO-3HA-RIαB was constructed by cloning RIα G325D into pAAV-hSyn-DIO-hM3D(Gq)-mCherry (Addgene, 44361). .. Addgene supplied pAAV-CaMKII-Cre (182736), and pAAV-Ple94-Cre (49125). pAAV-Dlx-Cre was constructed from pAAV-mDlx-NLS-mRuby2 (Addgene, 99130) by replacing mRuby2 with Cre. pCS2-3HA-SIK3 LxVPm2 was constructed by mutating two “LxVP” motifs to “AAAA” by site-directed mutagenesis. ..

    Article Title: PP2Acα regulates sleep amount and sleep homeostasis in mice
    Article Snippet: .. PP2Acα L309A was constructed by site-directed mutagenesis using specific primers that contain L309A mutations. pAAV-CaMKII-Cre (182736, Addgene) was purchased from Addgene. pAAV-mDlx-Cre was constructed by replacing mRuby2 with Cre from pAAV-mDlx-NLS-mRuby2 (99130, Addgene). ..

    Plasmid Preparation:

    Article Title: Paralog-specific intrabodies for PSD-93 and SAP102 expand the molecular toolkit to resolve excitatory synapse organization
    Article Snippet: For imaging, Xph binders were subcloned into pCAG_PSD95.FingR-eGFP-CCR5TC (gift from Don Arnold, USC, Addgene #46295) ( Gross et al .,2013 ) using KpnI and BglII restriction sites. .. Other fluorescent modules, mRuby2 (gift from Michael Lin, Addgene #40260) ( Lam et al .,2012 ), mNeonGreen (obtained by gene synthesis, Eurofins) ( Shaner et al .,2013 ), HaloTag (Promega, cat no G7971) and SNAPf (New England Biolabs, cat no N9183S) were next inserted in place of eGFP in the corresponding vector using BglII and NheI sites after an initial modification of the source vectors to introduce an NheI site between the fluorescent module and CCR5 ZF. .. AAV expressing vector containing Xph binders were subcloned into AAV-Syn-PSD95.FingR-eGFP-CCR5TC (gift from Xue Han, Addgene plasmid # 125693) ( Bensussen et al .,2020 ) by replacing the PSD95.FingR coding sequence using the NheI and SphI restriction sites.

    Article Title: Three-component CRISPR/Cas complex system and uses thereof
    Article Snippet: A fusion PCR of 5′ fragment of NLSPUFa with 3′ fragment of NLSPUb was used to create pAC1358:pmax-NLSPUFc_VP64. p65HSF1 activator ORF was amplified from MS2-P65-HSF1_GFP (Addgene: 61423) with FseI PacI sites to replace VP64 fragment in pAC164 to create pAC1410:pmax-dCas9_p65HSF1, and replace VP64 in pAC1355 and pAC1358 to create pAC1393: pmax-NLSPUFa_p65HSF1 and pAC1411:pmax-NLSPUFc_p65HSF1, respectively. .. Clover and mRuby2 were amplified from pcDNA3-Clover (Addgene #40259) and pcDNA3-mRuby2 (Addgene #40260) respectively with primers containing SgrAI and FseI cloning site, ligated with various FseI-PUF-PacI amplified from the above pAC1356 ̃1358 and vector digested from pAC149:pCR8-dCas9VP160 (Addgene #48221) to create gateway donor vectors pAC1402, pAC1403 and pAC1404 containing ORFs of Clover_PUFa and Clover_PUFc, mRuby2_PUFa, respectively. .. These ORFs are then transferred to PB3-neo vector by recombining with pAC1119:PB3-neo(−)-pmaxDEST(+) by LR Clonase (Invitrogen) to create expression vectors pAC1360 (Clover_PUFa), pAC1381 (Clover_PUFc) and pAC1362 (mRuby2_PUFa).

    Article Title: Linking molecular tension and cellular tractions: a multiscale approach to focal adhesion mechanics
    Article Snippet: Plasmid DNA expressing the vinculin tension sensor VinTS-tCRMod-GGSGGS7 (Addgene #111764), and its control tCRMod-GGSGGS7 (Addgene #111761), were gifts from Brenton Hoffman . .. Plasmid DNA expressing free Clover (Addgene #40259) and mRuby2 (Addgene #40260) were gifts from Michael Lin . .. Plasmid DNA containing the TRAF module (CTV, Addgene #27803) was a gift from Steven Vogel .

    Article Title: Linking molecular tension and cellular tractions: a multiscale approach to focal adhesion mechanics.
    Article Snippet: Plasmid DNA expressing the vinculin tension sensor VinTS-tCRMod-GGSGGS7 (Addgene #111764), and its control tCRMod-GGSGGS7 (Addgene #111761), were gifts from Brenton Hoffman. .. Plasmid DNA expressing free Clover (Addgene #40259) and mRuby2 (Addgene #40260) were gifts from Michael Lin. .. Plasmid DNA containing TRAF module (CTV, Addgene #27803) was a gift from Steven Vogel.

    Article Title: Linking molecular tension and cellular tractions: a multiscale approach to focal adhesion mechanics.
    Article Snippet: Plasmid DNA expressing the vinculin tension sensor VinTS-tCRMod-GGSGGS7 (Addgene #111764), and its control tCRMod-GGSGGS7 (Addgene #111761), were gifts from Brenton Hoffman30. .. Plasmid DNA expressing free Clover (Addgene #40259) and mRuby2 (Addgene #40260) were gifts from Michael Lin70. .. Plasmid DNA containing TRAF module (CTV, Addgene #27803) was a gift from Steven Vogel71.

    Modification:

    Article Title: Paralog-specific intrabodies for PSD-93 and SAP102 expand the molecular toolkit to resolve excitatory synapse organization
    Article Snippet: For imaging, Xph binders were subcloned into pCAG_PSD95.FingR-eGFP-CCR5TC (gift from Don Arnold, USC, Addgene #46295) ( Gross et al .,2013 ) using KpnI and BglII restriction sites. .. Other fluorescent modules, mRuby2 (gift from Michael Lin, Addgene #40260) ( Lam et al .,2012 ), mNeonGreen (obtained by gene synthesis, Eurofins) ( Shaner et al .,2013 ), HaloTag (Promega, cat no G7971) and SNAPf (New England Biolabs, cat no N9183S) were next inserted in place of eGFP in the corresponding vector using BglII and NheI sites after an initial modification of the source vectors to introduce an NheI site between the fluorescent module and CCR5 ZF. .. AAV expressing vector containing Xph binders were subcloned into AAV-Syn-PSD95.FingR-eGFP-CCR5TC (gift from Xue Han, Addgene plasmid # 125693) ( Bensussen et al .,2020 ) by replacing the PSD95.FingR coding sequence using the NheI and SphI restriction sites.

    Introduce:

    Article Title: Paralog-specific intrabodies for PSD-93 and SAP102 expand the molecular toolkit to resolve excitatory synapse organization
    Article Snippet: For imaging, Xph binders were subcloned into pCAG_PSD95.FingR-eGFP-CCR5TC (gift from Don Arnold, USC, Addgene #46295) ( Gross et al .,2013 ) using KpnI and BglII restriction sites. .. Other fluorescent modules, mRuby2 (gift from Michael Lin, Addgene #40260) ( Lam et al .,2012 ), mNeonGreen (obtained by gene synthesis, Eurofins) ( Shaner et al .,2013 ), HaloTag (Promega, cat no G7971) and SNAPf (New England Biolabs, cat no N9183S) were next inserted in place of eGFP in the corresponding vector using BglII and NheI sites after an initial modification of the source vectors to introduce an NheI site between the fluorescent module and CCR5 ZF. .. AAV expressing vector containing Xph binders were subcloned into AAV-Syn-PSD95.FingR-eGFP-CCR5TC (gift from Xue Han, Addgene plasmid # 125693) ( Bensussen et al .,2020 ) by replacing the PSD95.FingR coding sequence using the NheI and SphI restriction sites.

    Amplification:

    Article Title: Three-component CRISPR/Cas complex system and uses thereof
    Article Snippet: A fusion PCR of 5′ fragment of NLSPUFa with 3′ fragment of NLSPUb was used to create pAC1358:pmax-NLSPUFc_VP64. p65HSF1 activator ORF was amplified from MS2-P65-HSF1_GFP (Addgene: 61423) with FseI PacI sites to replace VP64 fragment in pAC164 to create pAC1410:pmax-dCas9_p65HSF1, and replace VP64 in pAC1355 and pAC1358 to create pAC1393: pmax-NLSPUFa_p65HSF1 and pAC1411:pmax-NLSPUFc_p65HSF1, respectively. .. Clover and mRuby2 were amplified from pcDNA3-Clover (Addgene #40259) and pcDNA3-mRuby2 (Addgene #40260) respectively with primers containing SgrAI and FseI cloning site, ligated with various FseI-PUF-PacI amplified from the above pAC1356 ̃1358 and vector digested from pAC149:pCR8-dCas9VP160 (Addgene #48221) to create gateway donor vectors pAC1402, pAC1403 and pAC1404 containing ORFs of Clover_PUFa and Clover_PUFc, mRuby2_PUFa, respectively. .. These ORFs are then transferred to PB3-neo vector by recombining with pAC1119:PB3-neo(−)-pmaxDEST(+) by LR Clonase (Invitrogen) to create expression vectors pAC1360 (Clover_PUFa), pAC1381 (Clover_PUFc) and pAC1362 (mRuby2_PUFa).

    Cloning:

    Article Title: Three-component CRISPR/Cas complex system and uses thereof
    Article Snippet: A fusion PCR of 5′ fragment of NLSPUFa with 3′ fragment of NLSPUb was used to create pAC1358:pmax-NLSPUFc_VP64. p65HSF1 activator ORF was amplified from MS2-P65-HSF1_GFP (Addgene: 61423) with FseI PacI sites to replace VP64 fragment in pAC164 to create pAC1410:pmax-dCas9_p65HSF1, and replace VP64 in pAC1355 and pAC1358 to create pAC1393: pmax-NLSPUFa_p65HSF1 and pAC1411:pmax-NLSPUFc_p65HSF1, respectively. .. Clover and mRuby2 were amplified from pcDNA3-Clover (Addgene #40259) and pcDNA3-mRuby2 (Addgene #40260) respectively with primers containing SgrAI and FseI cloning site, ligated with various FseI-PUF-PacI amplified from the above pAC1356 ̃1358 and vector digested from pAC149:pCR8-dCas9VP160 (Addgene #48221) to create gateway donor vectors pAC1402, pAC1403 and pAC1404 containing ORFs of Clover_PUFa and Clover_PUFc, mRuby2_PUFa, respectively. .. These ORFs are then transferred to PB3-neo vector by recombining with pAC1119:PB3-neo(−)-pmaxDEST(+) by LR Clonase (Invitrogen) to create expression vectors pAC1360 (Clover_PUFa), pAC1381 (Clover_PUFc) and pAC1362 (mRuby2_PUFa).

    Expressing:

    Article Title: Linking molecular tension and cellular tractions: a multiscale approach to focal adhesion mechanics
    Article Snippet: Plasmid DNA expressing the vinculin tension sensor VinTS-tCRMod-GGSGGS7 (Addgene #111764), and its control tCRMod-GGSGGS7 (Addgene #111761), were gifts from Brenton Hoffman . .. Plasmid DNA expressing free Clover (Addgene #40259) and mRuby2 (Addgene #40260) were gifts from Michael Lin . .. Plasmid DNA containing the TRAF module (CTV, Addgene #27803) was a gift from Steven Vogel .

    Article Title: Linking molecular tension and cellular tractions: a multiscale approach to focal adhesion mechanics.
    Article Snippet: Plasmid DNA expressing the vinculin tension sensor VinTS-tCRMod-GGSGGS7 (Addgene #111764), and its control tCRMod-GGSGGS7 (Addgene #111761), were gifts from Brenton Hoffman. .. Plasmid DNA expressing free Clover (Addgene #40259) and mRuby2 (Addgene #40260) were gifts from Michael Lin. .. Plasmid DNA containing TRAF module (CTV, Addgene #27803) was a gift from Steven Vogel.

    Article Title: Linking molecular tension and cellular tractions: a multiscale approach to focal adhesion mechanics.
    Article Snippet: Plasmid DNA expressing the vinculin tension sensor VinTS-tCRMod-GGSGGS7 (Addgene #111764), and its control tCRMod-GGSGGS7 (Addgene #111761), were gifts from Brenton Hoffman30. .. Plasmid DNA expressing free Clover (Addgene #40259) and mRuby2 (Addgene #40260) were gifts from Michael Lin70. .. Plasmid DNA containing TRAF module (CTV, Addgene #27803) was a gift from Steven Vogel71.



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    a Schematic overview of the experimental procedure: Day 1: preparation of thin (250 μm) and flat PAA gels; Day 2: surface functionalization with Sulfo-SANPAH and collagen coating in acidic solution; Day 3: seeding of cells transiently transfected with VinTS plasmid; Day 4: live cell imaging. b Imaging and data acquisition protocol. Step 1: stressed state bead and cell image acquisition (TFM imaging); Step 2: FLIM data acquisition of donor (Clover) channel (FRET imaging); Step3: intensity data of acceptor <t>(mRuby2)</t> channel; Step 4: relaxed state bead image acquired after the removal of the cells and the relaxation of the gel using a mild SDS solution. c Data processing workflow. Displacement fields generated from the overlay images of stressed and relaxed state bead images are calculated and subsequently converted into tractions. FRET efficiency is derived from the FRET trajectory between τ D and τ BG in the phasor space. FA structural and molecular properties are quantified from the acceptor intensity data after applying a binary FA mask. Illustrations were created using BioRender.com.
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    Image Search Results


    a Schematic overview of the experimental procedure: Day 1: preparation of thin (250 μm) and flat PAA gels; Day 2: surface functionalization with Sulfo-SANPAH and collagen coating in acidic solution; Day 3: seeding of cells transiently transfected with VinTS plasmid; Day 4: live cell imaging. b Imaging and data acquisition protocol. Step 1: stressed state bead and cell image acquisition (TFM imaging); Step 2: FLIM data acquisition of donor (Clover) channel (FRET imaging); Step3: intensity data of acceptor (mRuby2) channel; Step 4: relaxed state bead image acquired after the removal of the cells and the relaxation of the gel using a mild SDS solution. c Data processing workflow. Displacement fields generated from the overlay images of stressed and relaxed state bead images are calculated and subsequently converted into tractions. FRET efficiency is derived from the FRET trajectory between τ D and τ BG in the phasor space. FA structural and molecular properties are quantified from the acceptor intensity data after applying a binary FA mask. Illustrations were created using BioRender.com.

    Journal: Communications Biology

    Article Title: Linking molecular tension and cellular tractions: a multiscale approach to focal adhesion mechanics

    doi: 10.1038/s42003-026-09514-0

    Figure Lengend Snippet: a Schematic overview of the experimental procedure: Day 1: preparation of thin (250 μm) and flat PAA gels; Day 2: surface functionalization with Sulfo-SANPAH and collagen coating in acidic solution; Day 3: seeding of cells transiently transfected with VinTS plasmid; Day 4: live cell imaging. b Imaging and data acquisition protocol. Step 1: stressed state bead and cell image acquisition (TFM imaging); Step 2: FLIM data acquisition of donor (Clover) channel (FRET imaging); Step3: intensity data of acceptor (mRuby2) channel; Step 4: relaxed state bead image acquired after the removal of the cells and the relaxation of the gel using a mild SDS solution. c Data processing workflow. Displacement fields generated from the overlay images of stressed and relaxed state bead images are calculated and subsequently converted into tractions. FRET efficiency is derived from the FRET trajectory between τ D and τ BG in the phasor space. FA structural and molecular properties are quantified from the acceptor intensity data after applying a binary FA mask. Illustrations were created using BioRender.com.

    Article Snippet: Plasmid DNA expressing free Clover (Addgene #40259) and mRuby2 (Addgene #40260) were gifts from Michael Lin .

    Techniques: Transfection, Plasmid Preparation, Live Cell Imaging, Imaging, Generated, Derivative Assay

    a Traction and FRET overlay images of representative cells on soft (4.5 kPa) and stiff (13 kPa) PAA substrate. Traction arrows are scaled with traction magnitude. b Comparison of cell surface area between soft and stiff PAA substrate. n = 93 and 85 cells. c Comparison of cell-averaged tractions between the soft and stiff substrate ( n = 51 and 38 cells, respectively). Each data point represents a single cell. d Illustration of the structure and working principle of Tension Sensing Module (TSMod) during donor excitation; Clover (C) and mRuby2 (R) FRET pair is separated with a synthetic flexible polypeptide (GGSGGS) 7 ; FRET efficiency remains high due to the module’s inability to support tension. e Illustration of the structure and working principle of Vinculin Tension Sensor (VinTS) used; TSMod is inserted between vinculin head domain (VinHD) and vinculin tail domain (VinTD); FRET efficiency decreases as a result of applied tension. f Averaged focal adhesion FRET efficiencies per cell for VinTS and force-insensitive control TSMod, compared on three different substrates ( n = 20, 93, 29, 85, 22, and 26 cells). Each data point represents a single cell. The middle line of violin plots represents the median, and the upper and lower lines indicate the 75th and 25th percentiles. Statistical analysis between two groups were performed using non-parametric Mann–Whitney U test (Wilcoxon rank-sum test) for ( b , c ), and unpaired two-tailed parametric Student’s t test for ( f ): * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Illustrations ( d , e ) were created using BioRender.com.

    Journal: Communications Biology

    Article Title: Linking molecular tension and cellular tractions: a multiscale approach to focal adhesion mechanics

    doi: 10.1038/s42003-026-09514-0

    Figure Lengend Snippet: a Traction and FRET overlay images of representative cells on soft (4.5 kPa) and stiff (13 kPa) PAA substrate. Traction arrows are scaled with traction magnitude. b Comparison of cell surface area between soft and stiff PAA substrate. n = 93 and 85 cells. c Comparison of cell-averaged tractions between the soft and stiff substrate ( n = 51 and 38 cells, respectively). Each data point represents a single cell. d Illustration of the structure and working principle of Tension Sensing Module (TSMod) during donor excitation; Clover (C) and mRuby2 (R) FRET pair is separated with a synthetic flexible polypeptide (GGSGGS) 7 ; FRET efficiency remains high due to the module’s inability to support tension. e Illustration of the structure and working principle of Vinculin Tension Sensor (VinTS) used; TSMod is inserted between vinculin head domain (VinHD) and vinculin tail domain (VinTD); FRET efficiency decreases as a result of applied tension. f Averaged focal adhesion FRET efficiencies per cell for VinTS and force-insensitive control TSMod, compared on three different substrates ( n = 20, 93, 29, 85, 22, and 26 cells). Each data point represents a single cell. The middle line of violin plots represents the median, and the upper and lower lines indicate the 75th and 25th percentiles. Statistical analysis between two groups were performed using non-parametric Mann–Whitney U test (Wilcoxon rank-sum test) for ( b , c ), and unpaired two-tailed parametric Student’s t test for ( f ): * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Illustrations ( d , e ) were created using BioRender.com.

    Article Snippet: Plasmid DNA expressing free Clover (Addgene #40259) and mRuby2 (Addgene #40260) were gifts from Michael Lin .

    Techniques: Comparison, Single Cell, Control, MANN-WHITNEY, Two Tailed Test