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Exeter/17 July 2013
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'''Protocol:''' | '''Protocol:''' | ||
| - | 1 - Add 6 ul of pink co-precipitate to your nucleic acid sample and mix well for 30s. For samples that are greater than 200ul the pink co-precipitate can be increased accordingly but it isn't ever necessary to use over 20ul. | + | |
| - | 2 - Add an equal volume of SureClean to your nucleic acid solution and mix well. | + | '''1''' - Add 6 ul of pink co-precipitate to your nucleic acid sample and mix well for 30s. For samples that are greater than 200ul the pink co-precipitate can be increased accordingly but it isn't ever necessary to use over 20ul. |
| - | 3 - Centrifuge on max speed (!4,000 x g) for 10 mins. | + | |
| - | 4 - Carefully remove supernatant by pipette. | + | '''2''' - Add an equal volume of SureClean to your nucleic acid solution and mix well. |
| - | 5 - Add a volume of 70% ethanol that is equak to 2x the original sample volume and vortex for 10s. | + | |
| - | 6 - Repeat step 3. | + | '''3''' - Centrifuge on max speed (!4,000 x g) for 10 mins. |
| - | 7 - Remove supernatant and air dry to ensure all ethanol is removed. | + | |
| - | 8 - Resuspend pellet in the desired volume of TE, water or buffer. | + | '''4''' - Carefully remove supernatant by pipette. |
| - | 9 - Use NanoDrop to measure concentration of resulting purified sample - hopefully it will have improved! | + | |
| + | '''5''' - Add a volume of 70% ethanol that is equak to 2x the original sample volume and vortex for 10s. | ||
| + | |||
| + | '''6''' - Repeat step 3. | ||
| + | |||
| + | '''7''' - Remove supernatant and air dry to ensure all ethanol is removed. | ||
| + | |||
| + | '''8''' - Resuspend pellet in the desired volume of TE, water or buffer. | ||
| + | |||
| + | '''9''' - Use NanoDrop to measure concentration of resulting purified sample - hopefully it will have improved! | ||
Revision as of 15:54, 18 July 2013
MiniPreps of yesterday's liquid cultures
The plasmids were extracted from our liquid cultures, giving us DNA from...
- BBa_K592001, our green light sensor (CcaS)
- BBa_K592002, which codes for the protein (CcaR) which communicates between our green light sensor and inverter gene which eventually regulates transcription of the magenta pigment gene
- New Part 2, which has a promoter and RBS added to the FixJ coding region (BBa_K608002)
SureClean protocol
SureClean is a solution provided by bioline that allows column free nucleic acid purification. After a number of our digests we found the concentrations of resulting DNA to be unacceptably low when measured using the NanoDrop, fortunately SureClean provided us with a method of concentrating our samples.
Protocol:
1 - Add 6 ul of pink co-precipitate to your nucleic acid sample and mix well for 30s. For samples that are greater than 200ul the pink co-precipitate can be increased accordingly but it isn't ever necessary to use over 20ul.
2 - Add an equal volume of SureClean to your nucleic acid solution and mix well.
3 - Centrifuge on max speed (!4,000 x g) for 10 mins.
4 - Carefully remove supernatant by pipette.
5 - Add a volume of 70% ethanol that is equak to 2x the original sample volume and vortex for 10s.
6 - Repeat step 3.
7 - Remove supernatant and air dry to ensure all ethanol is removed.
8 - Resuspend pellet in the desired volume of TE, water or buffer.
9 - Use NanoDrop to measure concentration of resulting purified sample - hopefully it will have improved!
