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Exeter/11 July 2013
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| - | = | + | =Morning= |
| - | + | ||
| - | + | Today we will be trying to digest and ligate some of our BioBricks for the first time! | |
| - | + | ||
| - | + | '''MiniPrep''' | |
| - | + | ||
| - | + | We extracted the plasmids for... | |
| - | + | ||
| - | + | - BBa_K608002, a promoter and RBS | |
| - | + | - BBa_K592005, our FixJ protein intermediate | |
| + | - BBa_K592004, our blue light sensor | ||
| + | - BBa_B0015, a terminator | ||
| + | |||
| + | The normal MiniPrep protocol was followed, however the BBa_K608002 and BBa_B0015 Bricks only had 2/3rds as much genetic material collected as the other Bricks. (Plasmids collected from 2 Eppendorfs, as opposed to the usual 3) | ||
| + | |||
| + | =Digestions= | ||
| + | |||
| + | We are combining... | ||
| + | |||
| + | - a promoter and RBS to the blue light sensor (BBa_K608002 + BBa_K592004) | ||
| + | |||
| + | - a promoter and RBS to the FixJ protein coding region (BBa_K608002 + BBa_K592004) | ||
| + | |||
| + | - the magenta pigment to a terminator (BBa_K592012 + BBa_B0015) | ||
| + | |||
| + | - the yellow pigment to a terminator (BBa_K592010 + BBa_B0015) | ||
| + | |||
| + | First we have to work out the concentrations of each plasmid using a NanoDrop machine. | ||
| + | |||
| + | '''NanoDrop results''' | ||
| + | |||
| + | For our promoter and RBS Brick (BBa_K608002) and terminator Brick (BBa_B0015) we have two replicates. | ||
| + | |||
| + | - Promoter and RBS #1 - 29.1ng/ul | ||
| + | |||
| + | - Promoter and RBS #2 - 25.2ng/ul | ||
| + | |||
| + | - Terminator #1 - 27.6ng/ul | ||
| + | |||
| + | - Terminator #2 - 19.2ng/ul | ||
| + | |||
| + | - Blue light sensor - 38.3ng/ul | ||
| + | |||
| + | - FixJ intermediate - 58.1ng/ul | ||
| + | |||
| + | - Yellow pigment - 45.7ng/ul | ||
| + | |||
| + | - Magenta pigment - 72.0ng/ul | ||
| + | |||
| + | - RFP control - 43.4ng/ul | ||
=Ligate= | =Ligate= | ||
Revision as of 11:29, 16 July 2013
Morning
Today we will be trying to digest and ligate some of our BioBricks for the first time!
MiniPrep
We extracted the plasmids for...
- BBa_K608002, a promoter and RBS - BBa_K592005, our FixJ protein intermediate - BBa_K592004, our blue light sensor - BBa_B0015, a terminator
The normal MiniPrep protocol was followed, however the BBa_K608002 and BBa_B0015 Bricks only had 2/3rds as much genetic material collected as the other Bricks. (Plasmids collected from 2 Eppendorfs, as opposed to the usual 3)
Digestions
We are combining...
- a promoter and RBS to the blue light sensor (BBa_K608002 + BBa_K592004)
- a promoter and RBS to the FixJ protein coding region (BBa_K608002 + BBa_K592004)
- the magenta pigment to a terminator (BBa_K592012 + BBa_B0015)
- the yellow pigment to a terminator (BBa_K592010 + BBa_B0015)
First we have to work out the concentrations of each plasmid using a NanoDrop machine.
NanoDrop results
For our promoter and RBS Brick (BBa_K608002) and terminator Brick (BBa_B0015) we have two replicates.
- Promoter and RBS #1 - 29.1ng/ul
- Promoter and RBS #2 - 25.2ng/ul
- Terminator #1 - 27.6ng/ul
- Terminator #2 - 19.2ng/ul
- Blue light sensor - 38.3ng/ul
- FixJ intermediate - 58.1ng/ul
- Yellow pigment - 45.7ng/ul
- Magenta pigment - 72.0ng/ul
- RFP control - 43.4ng/ul
Ligate
- K592010 and B0015
- K592012 and B0015
- K592003 and Q04510
- K592005 and B0015
- K592002 and B0015
- K592001 and B0015
- K592004 and B0015
