"
Team:Macquarie Australia/WetLab/Protocols/PlasmidPreps
From 2014.igem.org
(Difference between revisions)
| Line 22: | Line 22: | ||
<li><a href="https://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/SDSPAGE">SDS-PAGE</a></li> | <li><a href="https://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/SDSPAGE">SDS-PAGE</a></li> | ||
<li><a href="https://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/Heat">Heat Shock</a></li> | <li><a href="https://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/Heat">Heat Shock</a></li> | ||
| + | <li><a href="https://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/Operons">Induction of Operons</a></li> | ||
</ul> | </ul> | ||
</div> | </div> | ||
Latest revision as of 13:10, 17 October 2014
Plasmid Prep
- Centrifuge @13,200 rpm for 10 min of 1.8mL of overnight cultures in 2mL Eppendorf’s
- Discard supernatant and add 1.9 mL of culture and centrifuge again @13,200 rpm for 10 min
- Re-suspend pelleted cells in P1 Buffer (250µl)
- Add 250µL of P2 buffer & invert 4-6 times (turns homogenous blue)
- Add 350µL of N3 & mix by inverting (turns colourless)
- Centrifuge @ 10min 13,000 rpm to obtain a pellet
- Transfer supernatant in QIA Prep Spin Column by pipetting
- Centrifuge 30-60 sec - discard flow through
- Wash QIA Prep Spin Column with 0.5mL of PB
- Centrifuge for 30-60 seconds - discard flow through
- Wash spin column by adding 0.75 mL PE buffer
- Centrifuge for 30-60 seconds - discard flow through and centrifuge for a further 1min
- Place QIAPrep Column in clean Eppendorf
- Elute DNA add 50µl of water, stand for 1min, centrifuge for 1min
- QIA prep - Spin miniprep buffer
Figure 1. Plasmid prep for QIAGEN QIAprep Spin Miniprep Kit.
