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e0554s camp glosensor assay promega  (New England Biolabs)


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    New England Biolabs e0554s camp glosensor assay promega
    E0554s Camp Glosensor Assay Promega, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 5716 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/glosensor+kit/Q5+Site-Directed+Mutagenesis+Kit/pm33571432-397-127-125
    Average 99 stars, based on 5716 article reviews
    e0554s camp glosensor assay promega - by Bioz Stars, 2026-10
    99/100 stars

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    Virus:

    Article Title: Ligand recognition and allosteric regulation of DRD1-Gs signaling complexes.
    Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial and virus strains E. coli cells DH10Bac Invitrogen 10361012 Chemicals, peptides, and recombinant proteins Benzamidine Sigma-Aldrich Cat# B6506 Leupeptin Sigma-Aldrich Cat# L2884 Lauryl Maltose Neopentyl Glycol (LMNG) Anatrace Cat# NG310 Glyco-diosgenin (GDN) Anatrace Cat# GDN101 Cholesterol hemisucinate TRIS Salt (CHS) Sigma-Aldrich Cat# C6512 Sodium Cholate Sigma-Aldrich Cat# C1254 TCEP Thermo Fisher Scientific Cat# 77720 Apyrase NEB Cat# M0398S Ethylenediamine Tetraacetic Acid (EDTA) Thermo Fisher Scientific Cat# S311-500 Monoclonal ANTI-FLAG M2-FITC antibody Sigma- Aldrich Cat# F4049 Fenoldopam Med Chem Express Cat# HY-B0735A A77636 TOCRIS Cat# 1701 Dopamine Selleck Cat# S2529 SKF83959 TOCRIS Cat# 2074 PW0464 This paper N/A LY3154207 This paper N/A [3H]-SCH23390 PerkinElmer NET930250UC Critical commercial assays HTRF Dynamic cAMP kit Cisbio Cat# 62AM4PEB Q5 site directed mutagenesis kit NEB Cat# E0554S cAMP Glosensor Assay Promega Cat# E1291 Deposited data Fenoldopam-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30392 PDB: 7CKW A77636-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30393 PDB: 7CKX PW0464-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30394 PDB: 7CKY Dopamine-PAM-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30395 PDB: 7CKZ SKF83959-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30452 PDB: 7CRH Experimental models: cell lines Insect cell line Sf9 Expression Systems N/A HEK293 cell ATCC N/A Primers for site-direct mutagenesis DRD1-S198A_forward: catctcaGCCtctgtaataagcttttacatccctg This paper N/A DRD1-S198A_reverse: attacagaGGCtgagatggcatatgtcctgctga This paper N/A DRD1-S202A_forward: CtgtaataGCCttttacatccctgtggccatcatg This paper N/A DRD1-S202A_reverse: tgtaaaaGGCtattacagaggatgagatggcatat This paper N/A DRD1-D103A_forward: cctttGCCatcatgtgctccactgcatccatc This paper N/A DRD1-D103A_reverse: agcacatgatGGCaaaggccacccagatg ttacagaag This paper N/A Recombinant DNA pFastBac1-HA-FLAG-DRD1-His*10 This paper N/A pcDNA3.1-HA-FLAG-DRD1 (WT and mutants) This paper N/A (Continued on next page) Cell 184, 943–956.e1–e6, February 18, 2021 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER pFastBac1-Gas This paper N/A pFastBac1Dual-Gbg This paper N/A Software and algorithms SerialEM Mastronarde DN https://bio3d.colorado.edu/SerialEM/ MotionCor2 (Zheng et al., 2017) https://emcore.ucsf.edu/ucsf-software Gctf Zhang, 2016 https://www.mrc-lmb.cam.ac.uk/ kzhang/Gctf/ Gautomatch Zhang, 2016 https://www.mrc-lmb.cam.ac.uk/kzhang/ Gautomatch Relion3.0 (Zivanov et al., 2018) https://www2.mrc-lmb.cam.ac.uk/relion CHIMERA Pettersen et al., 2004 http://www.cgl.ucsf.edu/chimera COOT (Emsley and Cowtan, 2004) https://www2.mrc-lmb.cam.ac.uk/ personal/pemsley/coot/ Phenix Adams et al., 2010 https://www.phenix-online.org PyMOL Schrodinger https://www.pymol.org/2/ Prism v.7.0 GraphPad Software https://www.graphpad.com Other Superdex 200 Increase column 10/300 GE healthcare Cat# 28990944 BIOTEK Cytation3 reader BIO-TEK N/A ESF921 cell culture medium Expression Systems Cat# 96-001 DMEM basic cell culture media GIBCO Cat# 11995 FBS (fetal bovine serum) Cell Box Cat# SAG-01U-02 Penicillin-Streptomycin GIBCO Cat# 15140122 Trypsin 0.5% EDTA GIBCO Cat# 25300054 9 cm2 Cell Culture Dish Corning Cat# 430166 6-well Cell Culture Dish Corning Cat# 430166 96-well Cell Culture Dish Corning Cat# 3599 96-well white microplate assay plates Corning Cat# 3917 ll Article

    Recombinant:

    Article Title: Ligand recognition and allosteric regulation of DRD1-Gs signaling complexes.
    Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial and virus strains E. coli cells DH10Bac Invitrogen 10361012 Chemicals, peptides, and recombinant proteins Benzamidine Sigma-Aldrich Cat# B6506 Leupeptin Sigma-Aldrich Cat# L2884 Lauryl Maltose Neopentyl Glycol (LMNG) Anatrace Cat# NG310 Glyco-diosgenin (GDN) Anatrace Cat# GDN101 Cholesterol hemisucinate TRIS Salt (CHS) Sigma-Aldrich Cat# C6512 Sodium Cholate Sigma-Aldrich Cat# C1254 TCEP Thermo Fisher Scientific Cat# 77720 Apyrase NEB Cat# M0398S Ethylenediamine Tetraacetic Acid (EDTA) Thermo Fisher Scientific Cat# S311-500 Monoclonal ANTI-FLAG M2-FITC antibody Sigma- Aldrich Cat# F4049 Fenoldopam Med Chem Express Cat# HY-B0735A A77636 TOCRIS Cat# 1701 Dopamine Selleck Cat# S2529 SKF83959 TOCRIS Cat# 2074 PW0464 This paper N/A LY3154207 This paper N/A [3H]-SCH23390 PerkinElmer NET930250UC Critical commercial assays HTRF Dynamic cAMP kit Cisbio Cat# 62AM4PEB Q5 site directed mutagenesis kit NEB Cat# E0554S cAMP Glosensor Assay Promega Cat# E1291 Deposited data Fenoldopam-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30392 PDB: 7CKW A77636-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30393 PDB: 7CKX PW0464-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30394 PDB: 7CKY Dopamine-PAM-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30395 PDB: 7CKZ SKF83959-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30452 PDB: 7CRH Experimental models: cell lines Insect cell line Sf9 Expression Systems N/A HEK293 cell ATCC N/A Primers for site-direct mutagenesis DRD1-S198A_forward: catctcaGCCtctgtaataagcttttacatccctg This paper N/A DRD1-S198A_reverse: attacagaGGCtgagatggcatatgtcctgctga This paper N/A DRD1-S202A_forward: CtgtaataGCCttttacatccctgtggccatcatg This paper N/A DRD1-S202A_reverse: tgtaaaaGGCtattacagaggatgagatggcatat This paper N/A DRD1-D103A_forward: cctttGCCatcatgtgctccactgcatccatc This paper N/A DRD1-D103A_reverse: agcacatgatGGCaaaggccacccagatg ttacagaag This paper N/A Recombinant DNA pFastBac1-HA-FLAG-DRD1-His*10 This paper N/A pcDNA3.1-HA-FLAG-DRD1 (WT and mutants) This paper N/A (Continued on next page) Cell 184, 943–956.e1–e6, February 18, 2021 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER pFastBac1-Gas This paper N/A pFastBac1Dual-Gbg This paper N/A Software and algorithms SerialEM Mastronarde DN https://bio3d.colorado.edu/SerialEM/ MotionCor2 (Zheng et al., 2017) https://emcore.ucsf.edu/ucsf-software Gctf Zhang, 2016 https://www.mrc-lmb.cam.ac.uk/ kzhang/Gctf/ Gautomatch Zhang, 2016 https://www.mrc-lmb.cam.ac.uk/kzhang/ Gautomatch Relion3.0 (Zivanov et al., 2018) https://www2.mrc-lmb.cam.ac.uk/relion CHIMERA Pettersen et al., 2004 http://www.cgl.ucsf.edu/chimera COOT (Emsley and Cowtan, 2004) https://www2.mrc-lmb.cam.ac.uk/ personal/pemsley/coot/ Phenix Adams et al., 2010 https://www.phenix-online.org PyMOL Schrodinger https://www.pymol.org/2/ Prism v.7.0 GraphPad Software https://www.graphpad.com Other Superdex 200 Increase column 10/300 GE healthcare Cat# 28990944 BIOTEK Cytation3 reader BIO-TEK N/A ESF921 cell culture medium Expression Systems Cat# 96-001 DMEM basic cell culture media GIBCO Cat# 11995 FBS (fetal bovine serum) Cell Box Cat# SAG-01U-02 Penicillin-Streptomycin GIBCO Cat# 15140122 Trypsin 0.5% EDTA GIBCO Cat# 25300054 9 cm2 Cell Culture Dish Corning Cat# 430166 6-well Cell Culture Dish Corning Cat# 430166 96-well Cell Culture Dish Corning Cat# 3599 96-well white microplate assay plates Corning Cat# 3917 ll Article

    Mutagenesis:

    Article Title: Ligand recognition and allosteric regulation of DRD1-Gs signaling complexes.
    Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Bacterial and virus strains E. coli cells DH10Bac Invitrogen 10361012 Chemicals, peptides, and recombinant proteins Benzamidine Sigma-Aldrich Cat# B6506 Leupeptin Sigma-Aldrich Cat# L2884 Lauryl Maltose Neopentyl Glycol (LMNG) Anatrace Cat# NG310 Glyco-diosgenin (GDN) Anatrace Cat# GDN101 Cholesterol hemisucinate TRIS Salt (CHS) Sigma-Aldrich Cat# C6512 Sodium Cholate Sigma-Aldrich Cat# C1254 TCEP Thermo Fisher Scientific Cat# 77720 Apyrase NEB Cat# M0398S Ethylenediamine Tetraacetic Acid (EDTA) Thermo Fisher Scientific Cat# S311-500 Monoclonal ANTI-FLAG M2-FITC antibody Sigma- Aldrich Cat# F4049 Fenoldopam Med Chem Express Cat# HY-B0735A A77636 TOCRIS Cat# 1701 Dopamine Selleck Cat# S2529 SKF83959 TOCRIS Cat# 2074 PW0464 This paper N/A LY3154207 This paper N/A [3H]-SCH23390 PerkinElmer NET930250UC Critical commercial assays HTRF Dynamic cAMP kit Cisbio Cat# 62AM4PEB Q5 site directed mutagenesis kit NEB Cat# E0554S cAMP Glosensor Assay Promega Cat# E1291 Deposited data Fenoldopam-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30392 PDB: 7CKW A77636-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30393 PDB: 7CKX PW0464-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30394 PDB: 7CKY Dopamine-PAM-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30395 PDB: 7CKZ SKF83959-DRD1-Gs EM map and coordinate This paper EMDB: EMD-30452 PDB: 7CRH Experimental models: cell lines Insect cell line Sf9 Expression Systems N/A HEK293 cell ATCC N/A Primers for site-direct mutagenesis DRD1-S198A_forward: catctcaGCCtctgtaataagcttttacatccctg This paper N/A DRD1-S198A_reverse: attacagaGGCtgagatggcatatgtcctgctga This paper N/A DRD1-S202A_forward: CtgtaataGCCttttacatccctgtggccatcatg This paper N/A DRD1-S202A_reverse: tgtaaaaGGCtattacagaggatgagatggcatat This paper N/A DRD1-D103A_forward: cctttGCCatcatgtgctccactgcatccatc This paper N/A DRD1-D103A_reverse: agcacatgatGGCaaaggccacccagatg ttacagaag This paper N/A Recombinant DNA pFastBac1-HA-FLAG-DRD1-His*10 This paper N/A pcDNA3.1-HA-FLAG-DRD1 (WT and mutants) This paper N/A (Continued on next page) Cell 184, 943–956.e1–e6, February 18, 2021 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER pFastBac1-Gas This paper N/A pFastBac1Dual-Gbg This paper N/A Software and algorithms SerialEM Mastronarde DN https://bio3d.colorado.edu/SerialEM/ MotionCor2 (Zheng et al., 2017) https://emcore.ucsf.edu/ucsf-software Gctf Zhang, 2016 https://www.mrc-lmb.cam.ac.uk/ kzhang/Gctf/ Gautomatch Zhang, 2016 https://www.mrc-lmb.cam.ac.uk/kzhang/ Gautomatch Relion3.0 (Zivanov et al., 2018) https://www2.mrc-lmb.cam.ac.uk/relion CHIMERA Pettersen et al., 2004 http://www.cgl.ucsf.edu/chimera COOT (Emsley and Cowtan, 2004) https://www2.mrc-lmb.cam.ac.uk/ personal/pemsley/coot/ Phenix Adams et al., 2010 https://www.phenix-online.org PyMOL Schrodinger https://www.pymol.org/2/ Prism v.7.0 GraphPad Software https://www.graphpad.com Other Superdex 200 Increase column 10/300 GE healthcare Cat# 28990944 BIOTEK Cytation3 reader BIO-TEK N/A ESF921 cell culture medium Expression Systems Cat# 96-001 DMEM basic cell culture media GIBCO Cat# 11995 FBS (fetal bovine serum) Cell Box Cat# SAG-01U-02 Penicillin-Streptomycin GIBCO Cat# 15140122 Trypsin 0.5% EDTA GIBCO Cat# 25300054 9 cm2 Cell Culture Dish Corning Cat# 430166 6-well Cell Culture Dish Corning Cat# 430166 96-well Cell Culture Dish Corning Cat# 3599 96-well white microplate assay plates Corning Cat# 3917 ll Article



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    FUT8 knockdown attenuates the FSH/FSHR signaling pathway. A, CCK8 cell proliferation assay. Cells were treated with 100 ng/mL rhFSH, and cell proliferation was measured by absorbance at a wavelength of 450 nm. Significant differences are shown as the means ± SD, n = 4. B, Immunofluorescence assay. KGN, KGN-KD and KGN-KD-Re cells were treated with FSH (400 ng/mL) and stained with anti-FSHR (red) (1:500) and DAPI (blue). C, Western blot. Cell lysates of KGN, KGN-KD and KGN-KD-Re cells were resolved by 7.5 % SDS-PAGE, transferred to a PVDF membrane and probed with anti-FSHR (1:2000). GAPDH was used as the reference gene for the cytoplasm. D, Western blot. Cell lysates of Fut8 +/+ and Fut8 −/− ovaries were resolved by 7.5 % SDS-PAGE, transferred to a PVDF membrane and probed with anti-FSHR (1:2000). GAPDH was used as a loading control. E, Quantitative real-time PCR. RNA was isolated from the granulosa cells of MOR and POR patients. The mRNA expression levels of FSHR were compared. The relative expression of mRNAs was normalized to GAPDH. Data are presented as the means ± SEM, n = 3. F, FSH-induced cAMP production was measured using a GloSensor cAMP Assay kit. Cells were seeded in a 6-well plate for 24 h, and then 100 μM of IBMX was added, to inhibit phosphodiesterases, for 20 min. The cAMP levels were measured in KGN, KGN-KD, and KGN-KD-Re cells following stimulation with 100 IU/L of rhFSH or 100 μM of forskolin for 5 min. Data are presented as the means ± SEM, n = 3. G, PKA-CREB signaling pathway and Akt-FoxO3a signaling pathway. Serum-starved KGN, KGN-KD, and KGN-KD-Re cells were treated with FSH for 30 min, and cell lysates were resolved by 10 % SDS-PAGE, followed by transfer to a PVDF membrane. Membranes were probed with anti-phospho-PKA (1:2000) and anti-PKA (1:2000); anti-phospho-CREB (1:2000) and anti-CREB (1:2000); and anti-KITL (1:2000); anti-phospho-Akt (1:2000) and anti-Akt (1:2000); anti-phospho-FoxO3a (1:2000) and anti-FoxO3a (1:2000), and GAPDH was used as a loading control. Quantitative data are represented as the means ± SD, n = 3. H, Western blot. Lysates of Fut8 +/+ and Fut8 −/− ovaries were resolved by 10 % SDS-PAGE, and probed with anti-KITL (1:2000), with GAPDH used as the loading control. Quantitative data are represented as the means ± SD, n = 3. I, Western blot. Lysates of Fut8 +/+ and Fut8 −/− ovaries were resolved by 10 % SDS-PAGE and probed with anti-pFoxO3a (1:2000) and anti-FoxO3a (1:2000). Quantitative data are represented as the means ± SD, n = 3. J, TUNEL assay in the Fut8 +/+ and Fut8 −/− ovarian follicles. Representative micrographs are shown. Quantitative data are represented as the means ± SD, n = 3. K, Western blot analysis. Cell lysates of Fut8 +/+ and Fut8 −/− ovaries were resolved by 7.5 % SDS-PAGE, transferred to a PVDF membrane, and probed with anti-BCL2 and anti-BAX (1:2000). Quantitative data are represented as the means ± SD, n = 3. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

    Journal: Journal of Advanced Research

    Article Title: Core fucosylation regulates the ovarian response via FSH receptor during follicular development

    doi: 10.1016/j.jare.2024.01.025

    Figure Lengend Snippet: FUT8 knockdown attenuates the FSH/FSHR signaling pathway. A, CCK8 cell proliferation assay. Cells were treated with 100 ng/mL rhFSH, and cell proliferation was measured by absorbance at a wavelength of 450 nm. Significant differences are shown as the means ± SD, n = 4. B, Immunofluorescence assay. KGN, KGN-KD and KGN-KD-Re cells were treated with FSH (400 ng/mL) and stained with anti-FSHR (red) (1:500) and DAPI (blue). C, Western blot. Cell lysates of KGN, KGN-KD and KGN-KD-Re cells were resolved by 7.5 % SDS-PAGE, transferred to a PVDF membrane and probed with anti-FSHR (1:2000). GAPDH was used as the reference gene for the cytoplasm. D, Western blot. Cell lysates of Fut8 +/+ and Fut8 −/− ovaries were resolved by 7.5 % SDS-PAGE, transferred to a PVDF membrane and probed with anti-FSHR (1:2000). GAPDH was used as a loading control. E, Quantitative real-time PCR. RNA was isolated from the granulosa cells of MOR and POR patients. The mRNA expression levels of FSHR were compared. The relative expression of mRNAs was normalized to GAPDH. Data are presented as the means ± SEM, n = 3. F, FSH-induced cAMP production was measured using a GloSensor cAMP Assay kit. Cells were seeded in a 6-well plate for 24 h, and then 100 μM of IBMX was added, to inhibit phosphodiesterases, for 20 min. The cAMP levels were measured in KGN, KGN-KD, and KGN-KD-Re cells following stimulation with 100 IU/L of rhFSH or 100 μM of forskolin for 5 min. Data are presented as the means ± SEM, n = 3. G, PKA-CREB signaling pathway and Akt-FoxO3a signaling pathway. Serum-starved KGN, KGN-KD, and KGN-KD-Re cells were treated with FSH for 30 min, and cell lysates were resolved by 10 % SDS-PAGE, followed by transfer to a PVDF membrane. Membranes were probed with anti-phospho-PKA (1:2000) and anti-PKA (1:2000); anti-phospho-CREB (1:2000) and anti-CREB (1:2000); and anti-KITL (1:2000); anti-phospho-Akt (1:2000) and anti-Akt (1:2000); anti-phospho-FoxO3a (1:2000) and anti-FoxO3a (1:2000), and GAPDH was used as a loading control. Quantitative data are represented as the means ± SD, n = 3. H, Western blot. Lysates of Fut8 +/+ and Fut8 −/− ovaries were resolved by 10 % SDS-PAGE, and probed with anti-KITL (1:2000), with GAPDH used as the loading control. Quantitative data are represented as the means ± SD, n = 3. I, Western blot. Lysates of Fut8 +/+ and Fut8 −/− ovaries were resolved by 10 % SDS-PAGE and probed with anti-pFoxO3a (1:2000) and anti-FoxO3a (1:2000). Quantitative data are represented as the means ± SD, n = 3. J, TUNEL assay in the Fut8 +/+ and Fut8 −/− ovarian follicles. Representative micrographs are shown. Quantitative data are represented as the means ± SD, n = 3. K, Western blot analysis. Cell lysates of Fut8 +/+ and Fut8 −/− ovaries were resolved by 7.5 % SDS-PAGE, transferred to a PVDF membrane, and probed with anti-BCL2 and anti-BAX (1:2000). Quantitative data are represented as the means ± SD, n = 3. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

    Article Snippet: FSH-induced cAMP production was measured using a GloSensor cAMP Assay kit (Promega).

    Techniques: Knockdown, Proliferation Assay, Immunofluorescence, Staining, Western Blot, SDS Page, Membrane, Control, Real-time Polymerase Chain Reaction, Isolation, Expressing, cAMP Assay, TUNEL Assay