Plasmid Preparation:Article Title: Cellular and biophysical barriers to lipid nanoparticle mediated delivery of RNA to the cytosol
Article Snippet: CHMP2A_GFP_N_term was a gift from Daniel Gerlich (Addgene plasmid # 31805). .. Lamp1-mScarlet-I (Addgene plasmid # 98827) and mScarletI-Rab7 (Addgene plasmid # 112955) were gifts from Dorus Gadella. mCherry-Rab4a-7 (Addgene plasmid # 55125) and mCherry-Rab11a-7 (Addgene plasmid # 55124) were gifts from Michael Davidson. mScarlet-EEA1 (Addgene plasmid # 169067) and GFP-Rab5B (Addgene plasmid # 61802) were gifts from Gia Voeltz. .. GFP-EEA1 wt was a gift from Silvia Corvera (Addgene plasmid # 42307).
Article Title: Cellular and biophysical barriers to lipid nanoparticle mediated delivery of RNA to the cytosol.
Article Snippet: Plasmids, siRNA, mRNA and primers CHMP2A_GFP_N_termwas a gift fromDaniel Gerlich (Addgene plasmid # 31805). .. Lamp1-mScarlet-I (Addgene plasmid # 98827) andmScarletIRab7 (Addgene plasmid # 112955) were gifts from Dorus Gadella. mCherry-Rab4a-7 (Addgene plasmid # 55125) and mCherry-Rab11a-7 (Addgene plasmid # 55124) were gifts from Michael Davidson. mScarlet-EEA1 (Addgene plasmid # 169067) and GFP-Rab5B (Addgene plasmid # 61802) were gifts from Gia Voeltz. .. GFP-EEA1 wt was a gift from Silvia Corvera (Addgene plasmid # 42307).
Article Title: Cellular and biophysical barriers to lipid nanoparticle mediated delivery of RNA to the cytosol
Article Snippet: CHMP2A_GFP_N_term was a gift from Daniel Gerlich (Addgene plasmid # 31805). .. Lamp1-mScarlet-I (Addgene plasmid # 98827) and mScarletI-Rab7 (Addgene plasmid # 112955) were gifts from Dorus Gadella. mCherry-Rab4a-7 (Addgene plasmid # 55125) and mCherry-Rab11a-7 (Addgene plasmid # 55124) were gifts from Michael Davidson. mScarlet-EEA1 (Addgene plasmid # 169067) and GFP-Rab5B (Addgene plasmid # 61802) were gifts from Gia Voeltz. .. GFP-EEA1 wt was a gift from Silvia Corvera (Addgene plasmid # 42307).
Construct:Article Title: Class A and B GPCRs trigger rapid Gα s translocation to late and slow recycling endosomes.
Article Snippet: .. pCMV-15F11-HA-mEGFP (Frankenbody) (#129590), pmRFP-C3-Rab5 (#14437), pmRFP-C3-Rab7 (#55125), mCherry-Rab11a-7 (#55124), and pEGFP-N1-Dyn1(K44A)-mRFP (#55795) constructs were purchased from Addgene. pCDNA3.1-3xHA-D1R and pCDNA3.1-HA-Gγ2 were purchased from UMR cDNA Resource Center (University of Missouri-Rolla, Rolla, USA). pRK5-myc-V2R, pcDNA3.1-G-Beta-1, pcDNA3.1-PTHR1, pSNAPf-HA-V2R, pcDNA3.1-Gαs-long-WT-RlucII(67), pcDNA3.1-Gαsshort-WT-RlucII(119), pcDNA3.1-CAAX-rGFP, pcDNA3.1-Tdr-rGFPRab4, pcDNA3.1-Tdr-rGFP-Rab5, pcDNA3.1-Tdr-rGFP-Rab7, pcDNA3.1-Tdr-rGFP-Rab11 and pcDNA3.1-GFP10-EPAC-RlucII, were kindly provided by Dr. Michel Bouvier (Université de Montréal, Montreal, Canada). pcDNA3-Flag-β2AR was a gift by Dr. Stéphane Laporte (McGill University, Montreal, Canada) and pmRFP-C3-Rab7 was a gift by Dr. Stéphane Lefrançois (Université deMontréal,Montreal, Canada). pSNAPfβ2AR was kindly provided by Dr. Roshanak Irannejad (University of California, San Francisco, USA). pcDNA3.1-Gαs(C3S)-EGFP and pcDNA3.1-Gαs(C3S) were generated by replacing cysteine with serine at position 3 of Gαs in the construct pcDNA3.1-Gαs-EGFP and pcDNA3.1- Communications Biology | (2025) 8:1715 11 Gαs-short, respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). pcDNA3.1-Gαs-EGFP and pcDNA3.1-Gαs-mini HA were generated with the primers described in Supplementary Table 1 using the Gibson Assembly cloning kit (New England Biolabs). pcDNA3.1-Gαs-EGFPwas based onHynes et al. with the exception that our construct is human insteadof rat39. pcDNA3.1-Myr-GαsEGFP and pcDNA3.1-Myr-Gαs-long-RLucII(67) were generated by adding a threonine at position 4 and a serine at position 6 of Gαs in the construct pcDNA3.1-Gαs-EGFP and pcDNA3.1-Gαs-long-RlucII(67), respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). .. Isoproterenol (#I6504-100MG), Epinephrine (#E4250), Salmeterol (#94749-08-3), Dobutamine (#D0676), Dopamine (#8502), Parathyroid hormone fragment human 1-34 (PTH1-34) (#P3796) and Anti-Flag rabbit (# F7425) were purchased from Sigma Aldrich (St. Louis, Missouri, USA).
Article Title: Class A and B GPCRs trigger rapid Gα s translocation to late and slow recycling endosomes
Article Snippet: .. pCMV-15F11-HA-mEGFP (Frankenbody) (#129590), pmRFP-C3-Rab5 (#14437), pmRFP-C3-Rab7 (#55125), mCherry-Rab11a-7 (#55124), and pEGFP-N1-Dyn1(K44A)-mRFP (#55795) constructs were purchased from Addgene. pCDNA3.1-3xHA-D R and pCDNA3.1-HA-Gγ2 were purchased from UMR cDNA Resource Center (University of Missouri-Rolla, Rolla, USA). pRK5-myc-V 2 R, pcDNA3.1-G-Beta-1, pcDNA3.1-PTHR1, pSNAPf-HA-V 2 R, pcDNA3.1-Gα s -long-WT-RlucII(67), pcDNA3.1-Gα s -short-WT-RlucII(119), pcDNA3.1-CAAX-rGFP, pcDNA3.1-Tdr-rGFP-Rab4, pcDNA3.1-Tdr-rGFP-Rab5, pcDNA3.1-Tdr-rGFP-Rab7, pcDNA3.1-Tdr-rGFP-Rab11 and pcDNA3.1-GFP10-EPAC-RlucII, were kindly provided by Dr. Michel Bouvier (Université de Montréal, Montreal, Canada). pcDNA3-Flag-β 2 AR was a gift by Dr. Stéphane Laporte (McGill University, Montreal, Canada) and pmRFP-C3-Rab7 was a gift by Dr. Stéphane Lefrançois (Université de Montréal, Montreal, Canada). pSNAPf-β 2 AR was kindly provided by Dr. Roshanak Irannejad (University of California, San Francisco, USA). pcDNA3.1-Gα s (C3S)-EGFP and pcDNA3.1-Gα s (C3S) were generated by replacing cysteine with serine at position 3 of Gα s in the construct pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -short, respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -mini HA were generated with the primers described in Supplementary Table using the Gibson Assembly cloning kit (New England Biolabs). pcDNA3.1-Gα s -EGFP was based on Hynes et al. with the exception that our construct is human instead of rat . pcDNA3.1-Myr-Gα s -EGFP and pcDNA3.1-Myr-Gα s -long-RLucII(67) were generated by adding a threonine at position 4 and a serine at position 6 of Gα s in the construct pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -long-RlucII(67), respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). .. Isoproterenol (#I6504-100MG), Epinephrine (#E4250), Salmeterol (#94749-08-3), Dobutamine (#D0676), Dopamine (#8502), Parathyroid hormone fragment human 1-34 (PTH1-34) (#P3796) and Anti-Flag rabbit (# F7425) were purchased from Sigma Aldrich (St. Louis, Missouri, USA).
Generated:Article Title: Class A and B GPCRs trigger rapid Gα s translocation to late and slow recycling endosomes.
Article Snippet: .. pCMV-15F11-HA-mEGFP (Frankenbody) (#129590), pmRFP-C3-Rab5 (#14437), pmRFP-C3-Rab7 (#55125), mCherry-Rab11a-7 (#55124), and pEGFP-N1-Dyn1(K44A)-mRFP (#55795) constructs were purchased from Addgene. pCDNA3.1-3xHA-D1R and pCDNA3.1-HA-Gγ2 were purchased from UMR cDNA Resource Center (University of Missouri-Rolla, Rolla, USA). pRK5-myc-V2R, pcDNA3.1-G-Beta-1, pcDNA3.1-PTHR1, pSNAPf-HA-V2R, pcDNA3.1-Gαs-long-WT-RlucII(67), pcDNA3.1-Gαsshort-WT-RlucII(119), pcDNA3.1-CAAX-rGFP, pcDNA3.1-Tdr-rGFPRab4, pcDNA3.1-Tdr-rGFP-Rab5, pcDNA3.1-Tdr-rGFP-Rab7, pcDNA3.1-Tdr-rGFP-Rab11 and pcDNA3.1-GFP10-EPAC-RlucII, were kindly provided by Dr. Michel Bouvier (Université de Montréal, Montreal, Canada). pcDNA3-Flag-β2AR was a gift by Dr. Stéphane Laporte (McGill University, Montreal, Canada) and pmRFP-C3-Rab7 was a gift by Dr. Stéphane Lefrançois (Université deMontréal,Montreal, Canada). pSNAPfβ2AR was kindly provided by Dr. Roshanak Irannejad (University of California, San Francisco, USA). pcDNA3.1-Gαs(C3S)-EGFP and pcDNA3.1-Gαs(C3S) were generated by replacing cysteine with serine at position 3 of Gαs in the construct pcDNA3.1-Gαs-EGFP and pcDNA3.1- Communications Biology | (2025) 8:1715 11 Gαs-short, respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). pcDNA3.1-Gαs-EGFP and pcDNA3.1-Gαs-mini HA were generated with the primers described in Supplementary Table 1 using the Gibson Assembly cloning kit (New England Biolabs). pcDNA3.1-Gαs-EGFPwas based onHynes et al. with the exception that our construct is human insteadof rat39. pcDNA3.1-Myr-GαsEGFP and pcDNA3.1-Myr-Gαs-long-RLucII(67) were generated by adding a threonine at position 4 and a serine at position 6 of Gαs in the construct pcDNA3.1-Gαs-EGFP and pcDNA3.1-Gαs-long-RlucII(67), respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). .. Isoproterenol (#I6504-100MG), Epinephrine (#E4250), Salmeterol (#94749-08-3), Dobutamine (#D0676), Dopamine (#8502), Parathyroid hormone fragment human 1-34 (PTH1-34) (#P3796) and Anti-Flag rabbit (# F7425) were purchased from Sigma Aldrich (St. Louis, Missouri, USA).
Article Title: Class A and B GPCRs trigger rapid Gα s translocation to late and slow recycling endosomes
Article Snippet: .. pCMV-15F11-HA-mEGFP (Frankenbody) (#129590), pmRFP-C3-Rab5 (#14437), pmRFP-C3-Rab7 (#55125), mCherry-Rab11a-7 (#55124), and pEGFP-N1-Dyn1(K44A)-mRFP (#55795) constructs were purchased from Addgene. pCDNA3.1-3xHA-D R and pCDNA3.1-HA-Gγ2 were purchased from UMR cDNA Resource Center (University of Missouri-Rolla, Rolla, USA). pRK5-myc-V 2 R, pcDNA3.1-G-Beta-1, pcDNA3.1-PTHR1, pSNAPf-HA-V 2 R, pcDNA3.1-Gα s -long-WT-RlucII(67), pcDNA3.1-Gα s -short-WT-RlucII(119), pcDNA3.1-CAAX-rGFP, pcDNA3.1-Tdr-rGFP-Rab4, pcDNA3.1-Tdr-rGFP-Rab5, pcDNA3.1-Tdr-rGFP-Rab7, pcDNA3.1-Tdr-rGFP-Rab11 and pcDNA3.1-GFP10-EPAC-RlucII, were kindly provided by Dr. Michel Bouvier (Université de Montréal, Montreal, Canada). pcDNA3-Flag-β 2 AR was a gift by Dr. Stéphane Laporte (McGill University, Montreal, Canada) and pmRFP-C3-Rab7 was a gift by Dr. Stéphane Lefrançois (Université de Montréal, Montreal, Canada). pSNAPf-β 2 AR was kindly provided by Dr. Roshanak Irannejad (University of California, San Francisco, USA). pcDNA3.1-Gα s (C3S)-EGFP and pcDNA3.1-Gα s (C3S) were generated by replacing cysteine with serine at position 3 of Gα s in the construct pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -short, respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -mini HA were generated with the primers described in Supplementary Table using the Gibson Assembly cloning kit (New England Biolabs). pcDNA3.1-Gα s -EGFP was based on Hynes et al. with the exception that our construct is human instead of rat . pcDNA3.1-Myr-Gα s -EGFP and pcDNA3.1-Myr-Gα s -long-RLucII(67) were generated by adding a threonine at position 4 and a serine at position 6 of Gα s in the construct pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -long-RlucII(67), respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). .. Isoproterenol (#I6504-100MG), Epinephrine (#E4250), Salmeterol (#94749-08-3), Dobutamine (#D0676), Dopamine (#8502), Parathyroid hormone fragment human 1-34 (PTH1-34) (#P3796) and Anti-Flag rabbit (# F7425) were purchased from Sigma Aldrich (St. Louis, Missouri, USA).
Mutagenesis:Article Title: Class A and B GPCRs trigger rapid Gα s translocation to late and slow recycling endosomes.
Article Snippet: .. pCMV-15F11-HA-mEGFP (Frankenbody) (#129590), pmRFP-C3-Rab5 (#14437), pmRFP-C3-Rab7 (#55125), mCherry-Rab11a-7 (#55124), and pEGFP-N1-Dyn1(K44A)-mRFP (#55795) constructs were purchased from Addgene. pCDNA3.1-3xHA-D1R and pCDNA3.1-HA-Gγ2 were purchased from UMR cDNA Resource Center (University of Missouri-Rolla, Rolla, USA). pRK5-myc-V2R, pcDNA3.1-G-Beta-1, pcDNA3.1-PTHR1, pSNAPf-HA-V2R, pcDNA3.1-Gαs-long-WT-RlucII(67), pcDNA3.1-Gαsshort-WT-RlucII(119), pcDNA3.1-CAAX-rGFP, pcDNA3.1-Tdr-rGFPRab4, pcDNA3.1-Tdr-rGFP-Rab5, pcDNA3.1-Tdr-rGFP-Rab7, pcDNA3.1-Tdr-rGFP-Rab11 and pcDNA3.1-GFP10-EPAC-RlucII, were kindly provided by Dr. Michel Bouvier (Université de Montréal, Montreal, Canada). pcDNA3-Flag-β2AR was a gift by Dr. Stéphane Laporte (McGill University, Montreal, Canada) and pmRFP-C3-Rab7 was a gift by Dr. Stéphane Lefrançois (Université deMontréal,Montreal, Canada). pSNAPfβ2AR was kindly provided by Dr. Roshanak Irannejad (University of California, San Francisco, USA). pcDNA3.1-Gαs(C3S)-EGFP and pcDNA3.1-Gαs(C3S) were generated by replacing cysteine with serine at position 3 of Gαs in the construct pcDNA3.1-Gαs-EGFP and pcDNA3.1- Communications Biology | (2025) 8:1715 11 Gαs-short, respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). pcDNA3.1-Gαs-EGFP and pcDNA3.1-Gαs-mini HA were generated with the primers described in Supplementary Table 1 using the Gibson Assembly cloning kit (New England Biolabs). pcDNA3.1-Gαs-EGFPwas based onHynes et al. with the exception that our construct is human insteadof rat39. pcDNA3.1-Myr-GαsEGFP and pcDNA3.1-Myr-Gαs-long-RLucII(67) were generated by adding a threonine at position 4 and a serine at position 6 of Gαs in the construct pcDNA3.1-Gαs-EGFP and pcDNA3.1-Gαs-long-RlucII(67), respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). .. Isoproterenol (#I6504-100MG), Epinephrine (#E4250), Salmeterol (#94749-08-3), Dobutamine (#D0676), Dopamine (#8502), Parathyroid hormone fragment human 1-34 (PTH1-34) (#P3796) and Anti-Flag rabbit (# F7425) were purchased from Sigma Aldrich (St. Louis, Missouri, USA).
Article Title: Class A and B GPCRs trigger rapid Gα s translocation to late and slow recycling endosomes
Article Snippet: .. pCMV-15F11-HA-mEGFP (Frankenbody) (#129590), pmRFP-C3-Rab5 (#14437), pmRFP-C3-Rab7 (#55125), mCherry-Rab11a-7 (#55124), and pEGFP-N1-Dyn1(K44A)-mRFP (#55795) constructs were purchased from Addgene. pCDNA3.1-3xHA-D R and pCDNA3.1-HA-Gγ2 were purchased from UMR cDNA Resource Center (University of Missouri-Rolla, Rolla, USA). pRK5-myc-V 2 R, pcDNA3.1-G-Beta-1, pcDNA3.1-PTHR1, pSNAPf-HA-V 2 R, pcDNA3.1-Gα s -long-WT-RlucII(67), pcDNA3.1-Gα s -short-WT-RlucII(119), pcDNA3.1-CAAX-rGFP, pcDNA3.1-Tdr-rGFP-Rab4, pcDNA3.1-Tdr-rGFP-Rab5, pcDNA3.1-Tdr-rGFP-Rab7, pcDNA3.1-Tdr-rGFP-Rab11 and pcDNA3.1-GFP10-EPAC-RlucII, were kindly provided by Dr. Michel Bouvier (Université de Montréal, Montreal, Canada). pcDNA3-Flag-β 2 AR was a gift by Dr. Stéphane Laporte (McGill University, Montreal, Canada) and pmRFP-C3-Rab7 was a gift by Dr. Stéphane Lefrançois (Université de Montréal, Montreal, Canada). pSNAPf-β 2 AR was kindly provided by Dr. Roshanak Irannejad (University of California, San Francisco, USA). pcDNA3.1-Gα s (C3S)-EGFP and pcDNA3.1-Gα s (C3S) were generated by replacing cysteine with serine at position 3 of Gα s in the construct pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -short, respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -mini HA were generated with the primers described in Supplementary Table using the Gibson Assembly cloning kit (New England Biolabs). pcDNA3.1-Gα s -EGFP was based on Hynes et al. with the exception that our construct is human instead of rat . pcDNA3.1-Myr-Gα s -EGFP and pcDNA3.1-Myr-Gα s -long-RLucII(67) were generated by adding a threonine at position 4 and a serine at position 6 of Gα s in the construct pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -long-RlucII(67), respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). .. Isoproterenol (#I6504-100MG), Epinephrine (#E4250), Salmeterol (#94749-08-3), Dobutamine (#D0676), Dopamine (#8502), Parathyroid hormone fragment human 1-34 (PTH1-34) (#P3796) and Anti-Flag rabbit (# F7425) were purchased from Sigma Aldrich (St. Louis, Missouri, USA).
Cloning:Article Title: Class A and B GPCRs trigger rapid Gα s translocation to late and slow recycling endosomes.
Article Snippet: .. pCMV-15F11-HA-mEGFP (Frankenbody) (#129590), pmRFP-C3-Rab5 (#14437), pmRFP-C3-Rab7 (#55125), mCherry-Rab11a-7 (#55124), and pEGFP-N1-Dyn1(K44A)-mRFP (#55795) constructs were purchased from Addgene. pCDNA3.1-3xHA-D1R and pCDNA3.1-HA-Gγ2 were purchased from UMR cDNA Resource Center (University of Missouri-Rolla, Rolla, USA). pRK5-myc-V2R, pcDNA3.1-G-Beta-1, pcDNA3.1-PTHR1, pSNAPf-HA-V2R, pcDNA3.1-Gαs-long-WT-RlucII(67), pcDNA3.1-Gαsshort-WT-RlucII(119), pcDNA3.1-CAAX-rGFP, pcDNA3.1-Tdr-rGFPRab4, pcDNA3.1-Tdr-rGFP-Rab5, pcDNA3.1-Tdr-rGFP-Rab7, pcDNA3.1-Tdr-rGFP-Rab11 and pcDNA3.1-GFP10-EPAC-RlucII, were kindly provided by Dr. Michel Bouvier (Université de Montréal, Montreal, Canada). pcDNA3-Flag-β2AR was a gift by Dr. Stéphane Laporte (McGill University, Montreal, Canada) and pmRFP-C3-Rab7 was a gift by Dr. Stéphane Lefrançois (Université deMontréal,Montreal, Canada). pSNAPfβ2AR was kindly provided by Dr. Roshanak Irannejad (University of California, San Francisco, USA). pcDNA3.1-Gαs(C3S)-EGFP and pcDNA3.1-Gαs(C3S) were generated by replacing cysteine with serine at position 3 of Gαs in the construct pcDNA3.1-Gαs-EGFP and pcDNA3.1- Communications Biology | (2025) 8:1715 11 Gαs-short, respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). pcDNA3.1-Gαs-EGFP and pcDNA3.1-Gαs-mini HA were generated with the primers described in Supplementary Table 1 using the Gibson Assembly cloning kit (New England Biolabs). pcDNA3.1-Gαs-EGFPwas based onHynes et al. with the exception that our construct is human insteadof rat39. pcDNA3.1-Myr-GαsEGFP and pcDNA3.1-Myr-Gαs-long-RLucII(67) were generated by adding a threonine at position 4 and a serine at position 6 of Gαs in the construct pcDNA3.1-Gαs-EGFP and pcDNA3.1-Gαs-long-RlucII(67), respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). .. Isoproterenol (#I6504-100MG), Epinephrine (#E4250), Salmeterol (#94749-08-3), Dobutamine (#D0676), Dopamine (#8502), Parathyroid hormone fragment human 1-34 (PTH1-34) (#P3796) and Anti-Flag rabbit (# F7425) were purchased from Sigma Aldrich (St. Louis, Missouri, USA).
Article Title: Class A and B GPCRs trigger rapid Gα s translocation to late and slow recycling endosomes
Article Snippet: .. pCMV-15F11-HA-mEGFP (Frankenbody) (#129590), pmRFP-C3-Rab5 (#14437), pmRFP-C3-Rab7 (#55125), mCherry-Rab11a-7 (#55124), and pEGFP-N1-Dyn1(K44A)-mRFP (#55795) constructs were purchased from Addgene. pCDNA3.1-3xHA-D R and pCDNA3.1-HA-Gγ2 were purchased from UMR cDNA Resource Center (University of Missouri-Rolla, Rolla, USA). pRK5-myc-V 2 R, pcDNA3.1-G-Beta-1, pcDNA3.1-PTHR1, pSNAPf-HA-V 2 R, pcDNA3.1-Gα s -long-WT-RlucII(67), pcDNA3.1-Gα s -short-WT-RlucII(119), pcDNA3.1-CAAX-rGFP, pcDNA3.1-Tdr-rGFP-Rab4, pcDNA3.1-Tdr-rGFP-Rab5, pcDNA3.1-Tdr-rGFP-Rab7, pcDNA3.1-Tdr-rGFP-Rab11 and pcDNA3.1-GFP10-EPAC-RlucII, were kindly provided by Dr. Michel Bouvier (Université de Montréal, Montreal, Canada). pcDNA3-Flag-β 2 AR was a gift by Dr. Stéphane Laporte (McGill University, Montreal, Canada) and pmRFP-C3-Rab7 was a gift by Dr. Stéphane Lefrançois (Université de Montréal, Montreal, Canada). pSNAPf-β 2 AR was kindly provided by Dr. Roshanak Irannejad (University of California, San Francisco, USA). pcDNA3.1-Gα s (C3S)-EGFP and pcDNA3.1-Gα s (C3S) were generated by replacing cysteine with serine at position 3 of Gα s in the construct pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -short, respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -mini HA were generated with the primers described in Supplementary Table using the Gibson Assembly cloning kit (New England Biolabs). pcDNA3.1-Gα s -EGFP was based on Hynes et al. with the exception that our construct is human instead of rat . pcDNA3.1-Myr-Gα s -EGFP and pcDNA3.1-Myr-Gα s -long-RLucII(67) were generated by adding a threonine at position 4 and a serine at position 6 of Gα s in the construct pcDNA3.1-Gα s -EGFP and pcDNA3.1-Gα s -long-RlucII(67), respectively, using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies). .. Isoproterenol (#I6504-100MG), Epinephrine (#E4250), Salmeterol (#94749-08-3), Dobutamine (#D0676), Dopamine (#8502), Parathyroid hormone fragment human 1-34 (PTH1-34) (#P3796) and Anti-Flag rabbit (# F7425) were purchased from Sigma Aldrich (St. Louis, Missouri, USA).
Recombinant:Article Title: Visualization of endogenous G proteins on endosomes and other organelles
Article Snippet: Recombinant DNA reagent , mCherry-Rab7a-7 , Addgene , 55127 , . .. Recombinant DNA reagent , mCherry-Rab11a-7 , Addgene , 55124 , . .. Recombinant DNA reagent , pmCherry-2xFYVE , Addgene , 140050 , .
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