From 2011.igem.org
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| - | '''Arabidopsis'''<br><br>
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| - | Seedling protocol
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| - | - Weigh in appr. 50mg of arabidopsis seeds in eppendorf tube (one tube per 250 ml erlenmayer flask) <br>
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| - | - Wash with 500 µl 70% EtOH for appr. 4-5 (or 2x2) minutes per tube (mix well) <br>
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| - | - Remove 70% EtOH and replace with 500µl 50% bleach (alternatively 5% Calcium hypochloride) <br>
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| - | - Incubate for 20 minutes <br>
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| - | - Wash several times with sterile ddH2O to remove bleach x3 <br>
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| - | - Vernalize seeds for 2-3 days <br><br>
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| - | Prepare sterile medium<br><br>
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| - | - Half strength Murashige salt (2.1g per liter ddH2O) <br>
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| - | - Add 0.5g MES salt (buffer) per liter of media <br>
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| - | - Adjust pH to 5.7-5.8 using 2M KOH <br>
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| - | - If required add 10g sucrose (e.g. 1% solution) – yes <br>
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| - | - Add 1% agarose = 10g/litre if making phytogel<br>
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| - | - Distribute into erlenmayer flasks (125 ml/250ml flask) <br>
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| - | - Autoclave for at least 15 minutes <br><br>
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| - | Some notes<br><br>
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| - | - Growth conditions : flasks on a shaker at appr. 200 rpm. Either under long day or constant light conditions <br>
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| - | - Grow seedlings for 5-6 days <br>
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| - | </html>
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Revision as of 10:43, 22 July 2011
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