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Team:HKU HongKong/Data/Protocols.html
From 2012.igem.org
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<li> | <li> | ||
| + | <p style="text-align: left"> | ||
<font color="#232323"> | <font color="#232323"> | ||
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<span style="font-size: 10pt; font-family: Tahoma; font-weight: 400">Place | <span style="font-size: 10pt; font-family: Tahoma; font-weight: 400">Place | ||
in water bath at 42°C for 90s.</span></font></li> | in water bath at 42°C for 90s.</span></font></li> | ||
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<span style="font-size: 10pt; font-family: Tahoma; font-weight: 400">Place | <span style="font-size: 10pt; font-family: Tahoma; font-weight: 400">Place | ||
| - | immediately back on ice for at least 2 min</span></font></li> | + | immediately back on ice for at least 2 min.</span></font></li> |
| + | <li> | ||
| + | <p class="MsoListParagraph" style="text-indent: -18.0pt; text-autospace: none; text-align: left; margin-left: 18.0pt"> | ||
| + | <font color="#232323"> | ||
| + | <span style="font-size: 10pt; font-family: Tahoma; font-weight: 400">Add | ||
| + | 800uL LB </span></font><font size="2" color="#232323"> | ||
| + | <span style="font-family: Tahoma; font-weight: 400">broth. Incubate for 1hr | ||
| + | at 37°C shaker</span></font><span style="font-size:11.0pt;font-family:"Times New Roman","serif"">.</span></li> | ||
| + | <li> | ||
| + | <p class="MsoListParagraph" style="text-indent: -18.0pt; text-autospace: none; text-align: left; margin-left: 18.0pt"> | ||
| + | <font size="2" color="#232323" face="Tahoma">Centrifuge at 130rpm for 5min | ||
| + | to remove supernatant. Re-suspend the pellet in about 10uL of supernatant.</font></li> | ||
| + | <li> | ||
| + | <p class="MsoListParagraph" style="text-indent: -18.0pt; text-autospace: none; text-align: left; margin-left: 18.0pt"> | ||
| + | <font face="Tahoma" size="2" color="#232323">Spread the entire re-suspended | ||
| + | pellet on ampicillin agar dishes.</font></li> | ||
| + | <li> | ||
| + | <p class="MsoListParagraph" style="text-indent: -18.0pt; text-autospace: none; text-align: left; margin-left: 18.0pt"> | ||
| + | <font face="Tahoma" size="2" color="#232323">Incubate for 12-16 hours at 37 | ||
| + | </font><font color="#232323"> | ||
| + | <span style="font-size: 10pt; font-family: Tahoma; font-weight: 400">°C.</span></font></p> | ||
| + | <p class="MsoListParagraph" style="text-indent: -18.0pt; text-autospace: none; text-align: left; margin-left: 18.0pt"> | ||
| + | </li> | ||
</ul> | </ul> | ||
<p class="MsoListParagraph" style="text-indent: -18.0pt; text-autospace: none; text-align: left; margin-left: 18.0pt"> </p> | <p class="MsoListParagraph" style="text-indent: -18.0pt; text-autospace: none; text-align: left; margin-left: 18.0pt"> </p> | ||
Revision as of 14:15, 23 September 2012
Team:HKU HK
From 2011.igem.org
Molecular Cloning Protocols
Transforming Competent E.coli with the
Plasmid:
We chose the biobrick K137076 to
ligate to the pvdQ gene for the following reasons:
-
Take out the competent E-coli cells from the -80 freezer. (Keep all tubes on ice).
-
Add 1uL of the plasmid DNA in 100uL competent cells (if from Kit). For transformation of ligated product, add 10uL of the ligated plasmid DNA in 100uL competent cells.
-
Incubate on ice for 10 min.
-
Place in water bath at 42°C for 90s.
-
Place immediately back on ice for at least 2 min.
-
Add 800uL LB broth. Incubate for 1hr at 37°C shaker.
-
Centrifuge at 130rpm for 5min to remove supernatant. Re-suspend the pellet in about 10uL of supernatant.
-
Spread the entire re-suspended pellet on ampicillin agar dishes.
-
Incubate for 12-16 hours at 37 °C.
