"
Exeter/17 July 2013
From 2013.igem.org
Fentwistle (Talk | contribs) |
GeorgiaDrew (Talk | contribs) |
||
| Line 8: | Line 8: | ||
- New Part 2, which has a promoter and RBS added to the FixJ coding region (BBa_K608002) | - 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! | ||
Revision as of 15:52, 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!
