Magnetic Beads:Article Title: Dual Sensing of Physiologic pH and Calcium by EFCAB9 Regulates Sperm Motility.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit polyclonal anti-mCatSper1 Laboratory of David E. Clapham; Ren et al., 2001 Cat#CatSper1; RRID:AB_2314097 Rabbit polyclonal anti-mCatSper2 Laboratory of David L. Garbers; Quill et al., 2001 N/A Rabbit polyclonal anti-mCatSper3 Laboratory of David E. Clapham; Qi et al., 2007 N/A Rabbit polyclonal anti-mCatSper4 Laboratory of David E. Clapham; Qi et al., 2007 N/A Rabbit polyclonal anti-mCatSperb Laboratory of David E. Clapham; Chung et al., 2011 N/A Rabbit polyclonal anti-mCatSperd Laboratory of David E. Clapham; Chung et al., 2011 N/A Rabbit polyclonal anti-mCatSperz Laboratory of Jean-Ju Chung; Chung et al., 2017 N/A Rabbit polyclonal anti-hCatSperz Laboratory of Jean-Ju Chung; Chung et al., 2017 N/A Rabbit polyclonal anti-mEFCAB9 This study N/A Rabbit polyclonal anti-GFP (FL) SantaCruz Cat# sc-8334; RRID:AB_641123 Mouse monoclonal anti-PMCA4 (clone JA9) Novus Biologicals Cat# NB300-569; RRID: AB_2061726 Mouse monoclonal anti-Flag (clone M2) Sigma-Aldrich Cat# F3165; RRID:AB_259529 Mouse monoclonal anti-acetylated tubulin Sigma-Aldrich Cat# 7451; RRID:AB_609894 Mouse monoclonal anti-Calmodulin Upstate Cat# 05-173; RRID:AB_309644 Mouse monoclonal anti-phosphotyrosine (clone 4G10) EMD Millipore Cat# 05-321; RRID:AB_309678 Mouse monoclonal anti-HA (clone 2-2.2.14) Pierce Cat# 26183; RRID:AB_2533056 Mouse monoclonal anti-MYC (clone 9E10) SantaCruz Cat# sc-40; RRID:AB_627268 Mouse monoclonal anti-HA (clone 6E2), HRP-conjugated CST Cat# 2999S; RRID:AB_1264166 Mouse monoclonal anti-MYC (clone 9B11), HRP-conjugated CST Cat# 2040S; RRID:AB_10707162 Mouse monoclonal anti-V5 (clone E10/V4RR), HRP-conjugated Invitrogen Cat# MA5-15253-HRP; RRID:AB_2537645 Goat anti-mouse IgG-HRP Jackson ImmunoResearch Cat# 115-035-003; RRID:AB_10015289 Goat anti-rabbit IgG-HRP Jackson ImmunoResearch Cat# 111-035-144; RRID:AB_2307391 Rat anti-mouse IgG Trueblot (clone eB144) Rockland Cat# 18-8817-31; RRID:AB_2610850 Anti-V5 Agarose Affinity Gel (clone V5-10) Sigma Aldrich Cat# A7345 Anti-HA Magnetic Beads (clone 2-2.2.14) Pierce Cat# 88836 Bacterial and Virus Strains NEB 10-beta Electrocompetent E. coli (High Efficiency) NEB Cat# C3019H BL21-CodonPlus (DE3)-RIL competent cells Agilent Technology Cat# 230245 Chemicals, Peptides, and Recombinant Proteins EmbryoMax M2 Medium (1X), Liquid, with phenol red EMD Millipore Cat# MR-015-D EmbryoMax Human Tubal Fluid (HTF) (1X), liquid, for Mouse IVF EMD Millipore Cat# MR-070-D (Continued on next page) e1 Cell 177, 1–15.e1–e9, May 30, 2019 .. REAGENT or RESOURCE SOURCE IDENTIFIER Mammalian expression vector for mouse CatSperd-V5 (pCAG-mCatSperd-V5) Laboratory of David E. Clapham; Chung et al., 2011 N/A Mammalian expression vector for human EFCAB9-HA (phCMV3-hEFCAB9-HA) This study N/A Mammalian expression vector for human CatSperz-V5 (phCMV3-hCatSpere-V5) This study N/A Mammalian expression vector for Flag-human CatSpere (phCMV3-Flag-hCatSpere) This study N/A Mammalian expression vector for Flag-human CatSperg (phCMV3-Flag-hCatSperg) This study N/A Mammalian bi-cystronical expression vector for human EFCAB9-HA amd human CatSperz-MYC (pcDNA3.1(-)-hEfcab9HA-P2A-hCatSperz-MYC) This study N/A Bactrerial expression vector for GSTmouse EFCAB9 (pGEX-6P2-mEfcab9) This study N/A Bactrerial expression vector for GSTmouse EFCAB9 (pET43a-mEfcab9) This study N/A Bactrerial expression vector for 6xHispET32a-gb1-mCatSperzEFCAB9 (pET32a-gb1-CatSperz) This study N/A Bactrerial expression vector for GSTmouse EFCAB92Mut (D72N/E160Q), (pGEX-6P2-mEfcab92Mut This study N/A Bactrerial expression vector for GSTmouse EFCAB94Mut (D72N/D82N and D149N/E160Q), (pGEX-6P2-mEfcab94Mut) This study N/A Bacterial expression vector for GSTmouse CatSper1 N terminus (1-150) (pGex-2T-mCatSper1-N150) Laboratory of David E. Clapham N/A Bacterial expression vector for mouse 6xHis-CatSper1 C terminus (574-686)-Flag (pET32a-mCatSper1-C-Flag) This study N/A Mouse Efcab9 ORF clone 6771929 Dharmacon Cat# MMM1013-202843024 Human Efcab9 ORF clone OHu00121D GenScript Cat# OHu00121D Human CatSpere ORF clone 5269307 Dharmacon Cat# MHS6278-202808312 Human CatSpere ORF clone 4823002 Dharmacon Cat# MHS1010-202742776 pGEX-6P2 GE Healthcare Cat# 28954650 pET43.1a(+) Novagen Cat# 70939 pET32a(+) Novagen Cat# 69015 Software and Algorithms Fiji Schindelin et al., 2012 http://fiji.sc Zen 2012 SP2 Carl Zeiss https://www.zeiss.com/microscopy/ us/products/microscope-software/ zen-lite.html SCA evolution Microptic https://www.micropticsl.com R R Foundation https://www.r-project.org/ MEGA6 Tamura et al., 2013 https://www.megasoftware.net/ Other Glutathione agarose Pierce Cat# 16100 HisPur Ni-NTA Resin Pierce Cat# 88221 Protein A/G PLUS-Agarose SantaCruz Cat# sc-2003 (Continued on next page) e3 Cell 177, 1–15.e1–e9, May 30, 2019 Please cite this article in press as: Hwang et al., Dual Sensing of Physiologic pH and Calcium by EFCAB9 Regulates Sperm Motility, Cell (2019), https://doi.org/10.1016/j.cell.2019.03.047
Virus:Article Title: Dual Sensing of Physiologic pH and Calcium by EFCAB9 Regulates Sperm Motility.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit polyclonal anti-mCatSper1 Laboratory of David E. Clapham; Ren et al., 2001 Cat#CatSper1; RRID:AB_2314097 Rabbit polyclonal anti-mCatSper2 Laboratory of David L. Garbers; Quill et al., 2001 N/A Rabbit polyclonal anti-mCatSper3 Laboratory of David E. Clapham; Qi et al., 2007 N/A Rabbit polyclonal anti-mCatSper4 Laboratory of David E. Clapham; Qi et al., 2007 N/A Rabbit polyclonal anti-mCatSperb Laboratory of David E. Clapham; Chung et al., 2011 N/A Rabbit polyclonal anti-mCatSperd Laboratory of David E. Clapham; Chung et al., 2011 N/A Rabbit polyclonal anti-mCatSperz Laboratory of Jean-Ju Chung; Chung et al., 2017 N/A Rabbit polyclonal anti-hCatSperz Laboratory of Jean-Ju Chung; Chung et al., 2017 N/A Rabbit polyclonal anti-mEFCAB9 This study N/A Rabbit polyclonal anti-GFP (FL) SantaCruz Cat# sc-8334; RRID:AB_641123 Mouse monoclonal anti-PMCA4 (clone JA9) Novus Biologicals Cat# NB300-569; RRID: AB_2061726 Mouse monoclonal anti-Flag (clone M2) Sigma-Aldrich Cat# F3165; RRID:AB_259529 Mouse monoclonal anti-acetylated tubulin Sigma-Aldrich Cat# 7451; RRID:AB_609894 Mouse monoclonal anti-Calmodulin Upstate Cat# 05-173; RRID:AB_309644 Mouse monoclonal anti-phosphotyrosine (clone 4G10) EMD Millipore Cat# 05-321; RRID:AB_309678 Mouse monoclonal anti-HA (clone 2-2.2.14) Pierce Cat# 26183; RRID:AB_2533056 Mouse monoclonal anti-MYC (clone 9E10) SantaCruz Cat# sc-40; RRID:AB_627268 Mouse monoclonal anti-HA (clone 6E2), HRP-conjugated CST Cat# 2999S; RRID:AB_1264166 Mouse monoclonal anti-MYC (clone 9B11), HRP-conjugated CST Cat# 2040S; RRID:AB_10707162 Mouse monoclonal anti-V5 (clone E10/V4RR), HRP-conjugated Invitrogen Cat# MA5-15253-HRP; RRID:AB_2537645 Goat anti-mouse IgG-HRP Jackson ImmunoResearch Cat# 115-035-003; RRID:AB_10015289 Goat anti-rabbit IgG-HRP Jackson ImmunoResearch Cat# 111-035-144; RRID:AB_2307391 Rat anti-mouse IgG Trueblot (clone eB144) Rockland Cat# 18-8817-31; RRID:AB_2610850 Anti-V5 Agarose Affinity Gel (clone V5-10) Sigma Aldrich Cat# A7345 Anti-HA Magnetic Beads (clone 2-2.2.14) Pierce Cat# 88836 Bacterial and Virus Strains NEB 10-beta Electrocompetent E. coli (High Efficiency) NEB Cat# C3019H BL21-CodonPlus (DE3)-RIL competent cells Agilent Technology Cat# 230245 Chemicals, Peptides, and Recombinant Proteins EmbryoMax M2 Medium (1X), Liquid, with phenol red EMD Millipore Cat# MR-015-D EmbryoMax Human Tubal Fluid (HTF) (1X), liquid, for Mouse IVF EMD Millipore Cat# MR-070-D (Continued on next page) e1 Cell 177, 1–15.e1–e9, May 30, 2019 .. REAGENT or RESOURCE SOURCE IDENTIFIER Mammalian expression vector for mouse CatSperd-V5 (pCAG-mCatSperd-V5) Laboratory of David E. Clapham; Chung et al., 2011 N/A Mammalian expression vector for human EFCAB9-HA (phCMV3-hEFCAB9-HA) This study N/A Mammalian expression vector for human CatSperz-V5 (phCMV3-hCatSpere-V5) This study N/A Mammalian expression vector for Flag-human CatSpere (phCMV3-Flag-hCatSpere) This study N/A Mammalian expression vector for Flag-human CatSperg (phCMV3-Flag-hCatSperg) This study N/A Mammalian bi-cystronical expression vector for human EFCAB9-HA amd human CatSperz-MYC (pcDNA3.1(-)-hEfcab9HA-P2A-hCatSperz-MYC) This study N/A Bactrerial expression vector for GSTmouse EFCAB9 (pGEX-6P2-mEfcab9) This study N/A Bactrerial expression vector for GSTmouse EFCAB9 (pET43a-mEfcab9) This study N/A Bactrerial expression vector for 6xHispET32a-gb1-mCatSperzEFCAB9 (pET32a-gb1-CatSperz) This study N/A Bactrerial expression vector for GSTmouse EFCAB92Mut (D72N/E160Q), (pGEX-6P2-mEfcab92Mut This study N/A Bactrerial expression vector for GSTmouse EFCAB94Mut (D72N/D82N and D149N/E160Q), (pGEX-6P2-mEfcab94Mut) This study N/A Bacterial expression vector for GSTmouse CatSper1 N terminus (1-150) (pGex-2T-mCatSper1-N150) Laboratory of David E. Clapham N/A Bacterial expression vector for mouse 6xHis-CatSper1 C terminus (574-686)-Flag (pET32a-mCatSper1-C-Flag) This study N/A Mouse Efcab9 ORF clone 6771929 Dharmacon Cat# MMM1013-202843024 Human Efcab9 ORF clone OHu00121D GenScript Cat# OHu00121D Human CatSpere ORF clone 5269307 Dharmacon Cat# MHS6278-202808312 Human CatSpere ORF clone 4823002 Dharmacon Cat# MHS1010-202742776 pGEX-6P2 GE Healthcare Cat# 28954650 pET43.1a(+) Novagen Cat# 70939 pET32a(+) Novagen Cat# 69015 Software and Algorithms Fiji Schindelin et al., 2012 http://fiji.sc Zen 2012 SP2 Carl Zeiss https://www.zeiss.com/microscopy/ us/products/microscope-software/ zen-lite.html SCA evolution Microptic https://www.micropticsl.com R R Foundation https://www.r-project.org/ MEGA6 Tamura et al., 2013 https://www.megasoftware.net/ Other Glutathione agarose Pierce Cat# 16100 HisPur Ni-NTA Resin Pierce Cat# 88221 Protein A/G PLUS-Agarose SantaCruz Cat# sc-2003 (Continued on next page) e3 Cell 177, 1–15.e1–e9, May 30, 2019 Please cite this article in press as: Hwang et al., Dual Sensing of Physiologic pH and Calcium by EFCAB9 Regulates Sperm Motility, Cell (2019), https://doi.org/10.1016/j.cell.2019.03.047
Recombinant:Article Title: Dual Sensing of Physiologic pH and Calcium by EFCAB9 Regulates Sperm Motility.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit polyclonal anti-mCatSper1 Laboratory of David E. Clapham; Ren et al., 2001 Cat#CatSper1; RRID:AB_2314097 Rabbit polyclonal anti-mCatSper2 Laboratory of David L. Garbers; Quill et al., 2001 N/A Rabbit polyclonal anti-mCatSper3 Laboratory of David E. Clapham; Qi et al., 2007 N/A Rabbit polyclonal anti-mCatSper4 Laboratory of David E. Clapham; Qi et al., 2007 N/A Rabbit polyclonal anti-mCatSperb Laboratory of David E. Clapham; Chung et al., 2011 N/A Rabbit polyclonal anti-mCatSperd Laboratory of David E. Clapham; Chung et al., 2011 N/A Rabbit polyclonal anti-mCatSperz Laboratory of Jean-Ju Chung; Chung et al., 2017 N/A Rabbit polyclonal anti-hCatSperz Laboratory of Jean-Ju Chung; Chung et al., 2017 N/A Rabbit polyclonal anti-mEFCAB9 This study N/A Rabbit polyclonal anti-GFP (FL) SantaCruz Cat# sc-8334; RRID:AB_641123 Mouse monoclonal anti-PMCA4 (clone JA9) Novus Biologicals Cat# NB300-569; RRID: AB_2061726 Mouse monoclonal anti-Flag (clone M2) Sigma-Aldrich Cat# F3165; RRID:AB_259529 Mouse monoclonal anti-acetylated tubulin Sigma-Aldrich Cat# 7451; RRID:AB_609894 Mouse monoclonal anti-Calmodulin Upstate Cat# 05-173; RRID:AB_309644 Mouse monoclonal anti-phosphotyrosine (clone 4G10) EMD Millipore Cat# 05-321; RRID:AB_309678 Mouse monoclonal anti-HA (clone 2-2.2.14) Pierce Cat# 26183; RRID:AB_2533056 Mouse monoclonal anti-MYC (clone 9E10) SantaCruz Cat# sc-40; RRID:AB_627268 Mouse monoclonal anti-HA (clone 6E2), HRP-conjugated CST Cat# 2999S; RRID:AB_1264166 Mouse monoclonal anti-MYC (clone 9B11), HRP-conjugated CST Cat# 2040S; RRID:AB_10707162 Mouse monoclonal anti-V5 (clone E10/V4RR), HRP-conjugated Invitrogen Cat# MA5-15253-HRP; RRID:AB_2537645 Goat anti-mouse IgG-HRP Jackson ImmunoResearch Cat# 115-035-003; RRID:AB_10015289 Goat anti-rabbit IgG-HRP Jackson ImmunoResearch Cat# 111-035-144; RRID:AB_2307391 Rat anti-mouse IgG Trueblot (clone eB144) Rockland Cat# 18-8817-31; RRID:AB_2610850 Anti-V5 Agarose Affinity Gel (clone V5-10) Sigma Aldrich Cat# A7345 Anti-HA Magnetic Beads (clone 2-2.2.14) Pierce Cat# 88836 Bacterial and Virus Strains NEB 10-beta Electrocompetent E. coli (High Efficiency) NEB Cat# C3019H BL21-CodonPlus (DE3)-RIL competent cells Agilent Technology Cat# 230245 Chemicals, Peptides, and Recombinant Proteins EmbryoMax M2 Medium (1X), Liquid, with phenol red EMD Millipore Cat# MR-015-D EmbryoMax Human Tubal Fluid (HTF) (1X), liquid, for Mouse IVF EMD Millipore Cat# MR-070-D (Continued on next page) e1 Cell 177, 1–15.e1–e9, May 30, 2019 .. REAGENT or RESOURCE SOURCE IDENTIFIER Mammalian expression vector for mouse CatSperd-V5 (pCAG-mCatSperd-V5) Laboratory of David E. Clapham; Chung et al., 2011 N/A Mammalian expression vector for human EFCAB9-HA (phCMV3-hEFCAB9-HA) This study N/A Mammalian expression vector for human CatSperz-V5 (phCMV3-hCatSpere-V5) This study N/A Mammalian expression vector for Flag-human CatSpere (phCMV3-Flag-hCatSpere) This study N/A Mammalian expression vector for Flag-human CatSperg (phCMV3-Flag-hCatSperg) This study N/A Mammalian bi-cystronical expression vector for human EFCAB9-HA amd human CatSperz-MYC (pcDNA3.1(-)-hEfcab9HA-P2A-hCatSperz-MYC) This study N/A Bactrerial expression vector for GSTmouse EFCAB9 (pGEX-6P2-mEfcab9) This study N/A Bactrerial expression vector for GSTmouse EFCAB9 (pET43a-mEfcab9) This study N/A Bactrerial expression vector for 6xHispET32a-gb1-mCatSperzEFCAB9 (pET32a-gb1-CatSperz) This study N/A Bactrerial expression vector for GSTmouse EFCAB92Mut (D72N/E160Q), (pGEX-6P2-mEfcab92Mut This study N/A Bactrerial expression vector for GSTmouse EFCAB94Mut (D72N/D82N and D149N/E160Q), (pGEX-6P2-mEfcab94Mut) This study N/A Bacterial expression vector for GSTmouse CatSper1 N terminus (1-150) (pGex-2T-mCatSper1-N150) Laboratory of David E. Clapham N/A Bacterial expression vector for mouse 6xHis-CatSper1 C terminus (574-686)-Flag (pET32a-mCatSper1-C-Flag) This study N/A Mouse Efcab9 ORF clone 6771929 Dharmacon Cat# MMM1013-202843024 Human Efcab9 ORF clone OHu00121D GenScript Cat# OHu00121D Human CatSpere ORF clone 5269307 Dharmacon Cat# MHS6278-202808312 Human CatSpere ORF clone 4823002 Dharmacon Cat# MHS1010-202742776 pGEX-6P2 GE Healthcare Cat# 28954650 pET43.1a(+) Novagen Cat# 70939 pET32a(+) Novagen Cat# 69015 Software and Algorithms Fiji Schindelin et al., 2012 http://fiji.sc Zen 2012 SP2 Carl Zeiss https://www.zeiss.com/microscopy/ us/products/microscope-software/ zen-lite.html SCA evolution Microptic https://www.micropticsl.com R R Foundation https://www.r-project.org/ MEGA6 Tamura et al., 2013 https://www.megasoftware.net/ Other Glutathione agarose Pierce Cat# 16100 HisPur Ni-NTA Resin Pierce Cat# 88221 Protein A/G PLUS-Agarose SantaCruz Cat# sc-2003 (Continued on next page) e3 Cell 177, 1–15.e1–e9, May 30, 2019 Please cite this article in press as: Hwang et al., Dual Sensing of Physiologic pH and Calcium by EFCAB9 Regulates Sperm Motility, Cell (2019), https://doi.org/10.1016/j.cell.2019.03.047
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