Team:UEA-JIC Norwich/Methods

From 2011.igem.org

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<a href="https://2011.igem.org/Team:UEA-JIC_Norwich/transformation"> High Efficiency Transformation Protocol </a>
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<a href="https://2011.igem.org/Team:UEA-JIC_Norwich/cultureprep"> Culture Preparation Protocol </a>
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<h1>Click on the happy scientist to open the protocol
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High Efficiency Transformation Protocol
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[[File:Happy_scientist.gif|High Efficiency Transformation Protocol|link=https://2011.igem.org/Team:UEA-JIC_Norwich/transformation]]
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<a href="https://2011.igem.org/Team:UEA-JIC_Norwich/glycerolstock"> Glycerol Stock Solution Protocol </a>
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Culture Preparation Protocol
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[[File:Happy_scientist.gif|Culture Preparation Protocol|link=https://2011.igem.org/Team:UEA-JIC_Norwich/cultureprep]]
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<a href="https://2011.igem.org/Team:UEA-JIC_Norwich/miniprep"> Qiagen Miniprep Protocol </a>
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Glycerol Stock Solution Protocol  
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[[File:Happy_scientist.gif|Glycerol Stock Solution Protocol|link=https://2011.igem.org/Team:UEA-JIC_Norwich/glycerolstock]]
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<a href="https://2011.igem.org/Team:UEA-JIC_Norwich/electrophoresis"> Gel Electrophoresis Protocol </a>
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Qiagen Miniprep Protocol
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[[File:Happy_scientist.gif|Qiagen Miniprep Protocol|link=https://2011.igem.org/Team:UEA-JIC_Norwich/miniprep]]
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<a href="https://2011.igem.org/Team:UEA-JIC_Norwich/pcr"> PCR protocol </a>
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Gel Electrophoresis Protocol
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[[File:Happy_scientist.gif|Gel Electrophoresis Protocol |link=https://2011.igem.org/Team:UEA-JIC_Norwich/electrophoresis]]
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<a href="https://2011.igem.org/Team:UEA-JIC_Norwich/pegtransformation"> PEG Transformation protocol </a>
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PCR Protocol
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[[File:Happy_scientist.gif|PCR Protocol |link=https://2011.igem.org/Team:UEA-JIC_Norwich/pcr]]
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<a href="https://2011.igem.org/Team:UEA-JIC_Norwich/algaeglassbeads"> Algae Transformation using glass beads protocol </a>
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PEG Transformation Protocol
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<p style="color:#000000">1. Transfer 50ml of algae culture into four 50ml screw top tubes.
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<p style="color:#000000">2. Place all four tubes in the eppendorf centrifuge at 800rpm for 9 minutes at 20-22°C.
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[[File:Happy_scientist.gif|PEG Transformation Protocol |link=https://2011.igem.org/Team:UEA-JIC_Norwich/pegtransformation]]
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<p style="color:#000000">3. Immediately pour contents of tubes into glass beaker and keep tubes upside down so no unwanted suspension mixes with the pellet.
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<p style="color:#000000">4. For all tubes resuspend cells in 750µl of TAP media.
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Algae Transformation using glass beads
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<p style="color:#000000">5. Wash pre prepared beads in ethanol removing excess gently.
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<p style="color:#000000">6. Wash beads approximately 2-3 times in autoclaved distilled water.
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[[File:Happy_scientist.gif|Algae Transformation using glass beads Protocol |link=https://2011.igem.org/Team:UEA-JIC_Norwich/algaeglassbeads]]
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<p style="color:#000000">7. Place approximately half the beads in a 1.5ml eppendorf tube for the control.
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<p style="color:#000000">8. Resuspend both tubes with 200µL of distilled water.
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Algae Transformation using electroporation
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<p style="color:#000000">9.
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<p style="color:#000000">10.
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[[File:Happy_scientist.gif|Algae Transformation using electroporation Protocol |link=https://2011.igem.org/Team:UEA-JIC_Norwich/algaeelectroporation]]
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<p style="color:#000000">11.
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<p style="color:#000000">12.
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Site Directed Mutagenesis Protocol
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[[File:Happy_scientist.gif|Site Directed Mutagenesis Protocol |link=https://2011.igem.org/Team:UEA-JIC_Norwich/sdm]]
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<p style="color:#000000">15.
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<p style="color:#000000">16.
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pGEM T-easy Vector Cloning protocol
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<p style="color:#000000">18.
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[[File:Happy_scientist.gif|pGEM T-easy Vector Cloning Protocol |link=https://2011.igem.org/Team:UEA-JIC_Norwich/vectorcloning]]
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<p style="color:#000000">19.
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<p style="color:#000000">20.
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Restriction Digest protocol
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[[File:Happy_scientist.gif|Restriction Digest Protocol|link=https://2011.igem.org/Team:UEA-JIC_Norwich/rdigest]]
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Poly "A" Tailing Protocol
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<h1 style="font-family:verdana;color:green">Algae Transformation Method using Electroporation</h1>
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<p style="color:#000000">1. Transfer 50ml of algae culture into four 50ml screw top tubes.
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[[File:Happy_scientist.gif|Poly "A" Tailing PCR Protocol|link=https://2011.igem.org/Team:UEA-JIC_Norwich/polyatailing]]
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<p style="color:#000000">2. Place all four tubes in the eppendorf centrifuge at 800rpm for 9 minutes at 20-22°C.
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<p style="color:#000000">3. Immediately pour contents of tubes into glass beaker and keep tubes upside down so no unwanted suspension mixes with the pellet.
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Ligation Protocol
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<p style="color:#000000">4. For all tubes resuspend cells in 250µl of TAP and sucrose media.
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<p style="color:#000000">5. Add linearised G-Luc plasmid (5µL)to the suspension and place on ice.
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[[File:Happy_scientist.gif|Ligation Protocol|link=https://2011.igem.org/Team:UEA-JIC_Norwich/ligation]]
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<p style="color:#000000">6. Transfer 250µL of algae suspension into 1mL cuvette.
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<p style="color:#000000">7. Subject cuvettes to electroporation at 2200v/ cm and place back on ice.
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Gel Extraction Protocol
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<p style="color:#000000">8. Add 500µL of starch solution to each cuvette and aspirate gently.
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<p style="color:#000000">9. Add 750µL of each sample onto an LB plate.
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[[File:Happy_scientist.gif|Gel Extraction Protocol|link=https://2011.igem.org/Team:UEA-JIC_Norwich/gelextract]]
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<p style="color:#000000">10.Parafilm the lid and plate to seal and place in a light incubator.
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<h1 style="font-family:verdana;color:green">Starch Preparation</h1>
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<p style="color:#000000">1. Wash starch powder with 10mL distilled water and resuspend.
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<p style="color:#000000">2. Centrifuge for five minutes at 4000rpm.
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<p style="color:#000000">3. Pour off supernatant.
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<p style="color:#000000">4. Resuspend with 500µL Ethanol.
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<p style="color:#000000">5. Centrifuge for five minutes at 4000rpm.
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<p style="color:#000000">6. Resuspend in 500µL TAP.
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<p style="color:#000000">7. Centrifuge for five minutes at 4000rpm, pour off supernatant, and repeat once.
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<p style="color:#000000">8. Resuspend in 500µL TAP with sucrose.
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Latest revision as of 21:18, 21 September 2011

University of East Anglia-JIC

UNIVERSITY OF EAST ANGLIA-JOHN INNES CENTRE

Research methodsbanner.jpg



Click on the happy scientist to open the protocol


High Efficiency Transformation Protocol
High Efficiency Transformation Protocol
Culture Preparation Protocol
Culture Preparation Protocol
Glycerol Stock Solution Protocol
Glycerol Stock Solution Protocol
Qiagen Miniprep Protocol
Qiagen Miniprep Protocol
Gel Electrophoresis Protocol
Gel Electrophoresis Protocol
PCR Protocol
PCR Protocol
PEG Transformation Protocol
PEG Transformation Protocol
Algae Transformation using glass beads
Algae Transformation using glass beads Protocol
Algae Transformation using electroporation
Algae Transformation using electroporation Protocol
Site Directed Mutagenesis Protocol
Site Directed Mutagenesis Protocol
pGEM T-easy Vector Cloning protocol
pGEM T-easy Vector Cloning Protocol
Restriction Digest protocol
Restriction Digest Protocol
Poly "A" Tailing Protocol
Poly "A" Tailing PCR Protocol
Ligation Protocol
Ligation Protocol
Gel Extraction Protocol

Gel Extraction Protocol

Retrieved from "http://2011.igem.org/Team:UEA-JIC_Norwich/Methods"