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Team:Hannover/Protocols/Transformation/Arabidopsis
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<h1><a href="https://2014.igem.org/Team:Hannover/Protocols" target="_blank">Protocols</a> / Floral Dipping</h1> | <h1><a href="https://2014.igem.org/Team:Hannover/Protocols" target="_blank">Protocols</a> / Floral Dipping</h1> | ||
| - | <table colspan="2" width=" | + | <table colspan="2" width="450px"><tr><td><h4>Material:</h4></td> |
<tr><td>Infiltration medium</td><td>2.2 g/L 0.5x MS medium,<br> 5% Saccharose,<br> 0.5 g/L MES,<br> 0.03% Silwett L-77, | <tr><td>Infiltration medium</td><td>2.2 g/L 0.5x MS medium,<br> 5% Saccharose,<br> 0.5 g/L MES,<br> 0.03% Silwett L-77, | ||
| - | pH 5.7 (adjust with 2N NaOH)</td><tr><tr><td>YEB medium</td><td>5 g/L Beef extract,<br> 1 g/L Yeast extract,<br> 5 g/L Peptone,<br> 5 g/l Sucrose,<br> 2 mM MgSO4</td></tr></table> | + | pH 5.7 (adjust with 2N NaOH)</td><tr><tr><td><td><td><td></tr><tr><td><td><td><td></tr><td>YEB medium</td><td>5 g/L Beef extract,<br> 1 g/L Yeast extract,<br> 5 g/L Peptone,<br> 5 g/l Sucrose,<br> 2 mM MgSO4</td></tr></table> |
<p><h4>Protocol:</h4></p> | <p><h4>Protocol:</h4></p> | ||
Revision as of 17:22, 15 October 2014
Protocols / Floral Dipping
Material: |
|||
| Infiltration medium | 2.2 g/L 0.5x MS medium, 5% Saccharose, 0.5 g/L MES, 0.03% Silwett L-77, pH 5.7 (adjust with 2N NaOH) | ||
| YEB medium | 5 g/L Beef extract, 1 g/L Yeast extract, 5 g/L Peptone, 5 g/l Sucrose, 2 mM MgSO4 |
Protocol:
Cultivate R. radiobacter cells in 5 ml antbiotics containing YEB media for 2 days at 28 °C. To start your main culture, add 1 ml of that suspension to 50 ml of antibiotics containing YEB medium and incubate it for 1 d at 28 °C. When the cell culture reached an optical density (OD600) of ~1.0, the bacteria were placed on ice and centrifuged for 15 min at 3000 rpm and 4 °C. While the supernatant was decanted, the precipitate was dissolved in 200 ml infiltration medium. The Inflorescence (mind. 15 cm) of Arabidopsis plants were dipped in the solution for 6 min. To minimize contaminations, remove already opened flowers and seed containing siliques. Place the pot in a horizontal position on a tray and cover all plants in plastic bags . The bags can be removed at the next day. .
