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Team:Bordeaux/NotebookJune2
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<h2>2<sup>nd</sup> June </h2> | <h2>2<sup>nd</sup> June </h2> | ||
| - | < | + | <div class="type-exp"> Molecular biology</div> |
<ul> | <ul> | ||
| - | <li>< | + | <li><div>We use DH5α E.coli for cloning and others manipulations</div></li> |
| - | <li> < | + | <li> <div class="title1">Terrific Broth : 1L</div></li> |
| - | + | ||
<table> | <table> | ||
<tr><td>Deionized H2O</td><td>to 900 mL</td></tr> | <tr><td>Deionized H2O</td><td>to 900 mL</td></tr> | ||
| Line 13: | Line 13: | ||
<tr><td>Yeast extract</td><td>24 g</td></tr> | <tr><td>Yeast extract</td><td>24 g</td></tr> | ||
<tr><td>Glycerol 10%</td><td>4 mL</td></tr> | <tr><td>Glycerol 10%</td><td>4 mL</td></tr> | ||
| - | </table | + | </table> |
<br> | <br> | ||
| - | < | + | <div>Shake until the solutes have dissolved and then sterilize by autoclaving for 20 min at 15 psi (1.05 kg/cm2) on liquid cycle. Allow the solution to cool to 60°C or less, and then add 100 mL of a sterile solution of 0.17 M KH2PO4, 0.72 M K2HPO4.</div> |
| - | < | + | <div>(This solution is made by dissolving 2.31 g of KH2PO4 and 12.54 g of K2HPO4 in 90 mL of H2O. After the salts have dissolved, adjust the volume of the solution to 100 mL with H2O and sterilize by autoclaving for 20 min at 15 psi [1.05 kg/cm2] on liquid cycle.)</div> |
| - | <li>< | + | <li><div class="title1">SOB : 1L</div></li> |
| - | + | <table> | |
<tr><td>2%</td><td>Bactotryptone</td><td>20g</td></tr> | <tr><td>2%</td><td>Bactotryptone</td><td>20g</td></tr> | ||
<tr><td>0,5</td><td>Yeast Extract</td><td>5g</td></tr> | <tr><td>0,5</td><td>Yeast Extract</td><td>5g</td></tr> | ||
| Line 25: | Line 25: | ||
<tr><td>10mM</td><td>MgCl2</td><td>10mL (1M stock)</td></tr> | <tr><td>10mM</td><td>MgCl2</td><td>10mL (1M stock)</td></tr> | ||
<tr><td>10mM</td><td>MgSO4</td><td>10mL (1M stock)</td></tr> | <tr><td>10mM</td><td>MgSO4</td><td>10mL (1M stock)</td></tr> | ||
| - | </table | + | </table> |
| - | < | + | <div>pH media to 7 with NaOH, then autoclave.</div> |
| - | < | + | <div class="title1">Transformation Broth (TB): 1L </div> |
| - | + | <table> | |
<tr><td>10 mM</td><td>Pipes</td><td>3.02 g</td></tr> | <tr><td>10 mM</td><td>Pipes</td><td>3.02 g</td></tr> | ||
<tr><td>15 mM</td><td>CaCl2</td><td>2.21 g</td></tr> | <tr><td>15 mM</td><td>CaCl2</td><td>2.21 g</td></tr> | ||
<tr><td>250 mM</td><td>KCl</td><td>18.64 g</td></tr> | <tr><td>250 mM</td><td>KCl</td><td>18.64 g</td></tr> | ||
| - | </table | + | </table> |
| - | < | + | <div>Mix Pipes, CaCl2 and KCl in 900mL of Millipore water. Add NaOH until pH 6.7 (Solution become uncolored). Add MnCl2 (See below), stir, adjust volume to 1L, then filter sterilize. Store at 4°C.</div> |
| - | + | <table> | |
<tr><td>55 mM</td><td>MnCl2</td><td>10.89 g</td></tr> | <tr><td>55 mM</td><td>MnCl2</td><td>10.89 g</td></tr> | ||
| - | </table | + | </table> |
| - | < | + | <div> <b>DAY ONE :</b> Grow 12 mL overnight culture of favorite strain of E.Coli in 2XTY (~LB) (We use a select gold Efficiency, which are comparable to DH5α according to Bioline). Make SOB and TB.</div> |
</ul> | </ul> | ||
</body> | </body> | ||
</html> | </html> | ||
Revision as of 17:56, 17 October 2014
2nd June
Molecular biology
- We use DH5α E.coli for cloning and others manipulations
- Terrific Broth : 1L
- SOB : 1L
| Deionized H2O | to 900 mL |
| Bactotryptone | 12 g |
| Yeast extract | 24 g |
| Glycerol 10% | 4 mL |
Shake until the solutes have dissolved and then sterilize by autoclaving for 20 min at 15 psi (1.05 kg/cm2) on liquid cycle. Allow the solution to cool to 60°C or less, and then add 100 mL of a sterile solution of 0.17 M KH2PO4, 0.72 M K2HPO4.
(This solution is made by dissolving 2.31 g of KH2PO4 and 12.54 g of K2HPO4 in 90 mL of H2O. After the salts have dissolved, adjust the volume of the solution to 100 mL with H2O and sterilize by autoclaving for 20 min at 15 psi [1.05 kg/cm2] on liquid cycle.)
| 2% | Bactotryptone | 20g |
| 0,5 | Yeast Extract | 5g |
| 10mM | NaCl | 2mL (5M stock) |
| 2.5mM | KCl | 2.5mL (1M stock) |
| 10mM | MgCl2 | 10mL (1M stock) |
| 10mM | MgSO4 | 10mL (1M stock) |
pH media to 7 with NaOH, then autoclave.
Transformation Broth (TB): 1L
| 10 mM | Pipes | 3.02 g |
| 15 mM | CaCl2 | 2.21 g |
| 250 mM | KCl | 18.64 g |
Mix Pipes, CaCl2 and KCl in 900mL of Millipore water. Add NaOH until pH 6.7 (Solution become uncolored). Add MnCl2 (See below), stir, adjust volume to 1L, then filter sterilize. Store at 4°C.
| 55 mM | MnCl2 | 10.89 g |
DAY ONE : Grow 12 mL overnight culture of favorite strain of E.Coli in 2XTY (~LB) (We use a select gold Efficiency, which are comparable to DH5α according to Bioline). Make SOB and TB.
