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[DNA Library] Phenol/Freeze RNA Prep (Yeast) 日期：2005-07-18 00:00:00 点击：159 好评：0
Tuesday, November 18, 2003 Description Phenol/Freeze RNA Prep (Yeast) Procedure 1. Grow 10 ml of yeast to 5E6 cells/ml. 2. Harvest cells by centrifugation and resuspend cells in 400 l of AE buffer. 3. Transfer cells to a 1.5 ml microcentri...
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[DNA Library] Gene Disruption via PCR (Yeast) 日期：2005-07-18 00:00:00 点击：191 好评：0
Tuesday, November 18, 2003 Description Gene Disruption via PCR (Yeast) Procedure Reaction mix: 5 l 10X Taq Buffer 5 l 25 mM MgCl2 2 l 10 mM dNTP's 10-100 ng template DNA 25 pmols of each primer 0.5 l Taq polymerase (2 Units) ______________...
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[DNA Library] Electroporation (Yeast) 日期：2005-07-18 00:00:00 点击：167 好评：0
Tuesday, November 18, 2003 Description Electroporation (Yeast) Procedure 1. Inoculate 50ml YEPD with a colony and grow with shaking at 30 deg C until early stationary (~0.6-2E8 cells/ml). 2. Harvest in a 50ml sterile conical tube in GPR ce...
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[DNA Library] PLATE Transformation 日期：2005-07-18 00:00:00 点击：132 好评：0
Tuesday, November 18, 2003 Description PLATE Transformation Procedure COMPETENCY: Inoculate a single colony into 100 ml YPD (or selective media if necessary). Grow on shaker to O.D.(600) = 1.0 (O.D.(600) between 0.6 and 1.8 is fine) at 30o...
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[DNA Library] Quick transformation 日期：2005-07-18 00:00:00 点击：107 好评：0
Tuesday, November 18, 2003 Description Quick transformation Procedure Take 0.5 ml of culture and spin 10 sec in microfuge. Decant the tube by inverting it and shaking it once. Alternatively, one can pick a colony (2-3 mm in diameter) from...
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[DNA Library] High efficiency LiAc transformation 日期：2005-07-18 00:00:00 点击：137 好评：0
Tuesday, November 18, 2003 Description High efficiency LiAc transformation Procedure 1. Inoculate 2-5 mls of liquid YPED or 10 ml synthetic media and incubate with shaking overnight at 30o C. 2. Count o/n culture and inoculate 50 mls of wa...
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[DNA Library] Colony Plasmid Rescue 日期：2005-07-18 00:00:00 点击：169 好评：0
Tuesday, November 18, 2003 Description Colony Plasmid Rescue Procedure Start with a fresh plate of yeast, with large colonies grown for 3-4 days. Prepare eppendorfs with 20 l aqueous buffer (2% Triton X-100, 1% SDS, 100 mM NaCl, 10 mM Tris...
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[DNA Library] High Efficency Yeast Plasmid Rescue 日期：2005-07-18 00:00:00 点击：106 好评：0
Tuesday, November 18, 2003 Description High Efficency Yeast Plasmid Rescue Procedure Grow yeast overnight. Spin down 1.5 ml of culture in microfuge tubes. Decant Add 250 l of Qiagen buffer P1 to each tube Add about 250 l of glass beads Bea...
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[DNA Library] Making electrocompetent Agrobacterium 日期：2005-07-18 00:00:00 点击：143 好评：0
Monday, November 10, 2003 Description Making electrocompetent Agrobacterium Procedure 1. Start a 5ml culture (LB plus appropriate selection - for GV2260 this is Rif25, Strep50 and Carb50) from a freshly streaked plate. Incubate at 28C over...
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[DNA Library] Electroporation of A. tumfaciens cells 日期：2005-07-18 00:00:00 点击：118 好评：0
Monday, November 10, 2003 Description Electroporation of A. tum faciens cells Procedure 1. Make a overnight culture of the A.tum. strain at 28 C. 2. Dilute cefis 10 times in the desired medium (8 ml for 1 etectroporation). 3. Grow cells at...