Team:TorontoMaRSDiscovery/Notebook/October
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+ | =October 1, 2009= | ||
+ | #Miniprepped Tet, 5, EncY, C | ||
+ | #Ran minipreps and K plasmid on a gel | ||
+ | #*Besides Tet, all samples looked fine | ||
+ | #Poured K plates | ||
+ | |||
+ | =October 2, 2009= | ||
+ | #Digested K, EncY and ran on gel | ||
+ | |||
+ | =October 3, 2009= | ||
+ | #Digested EncY and old Tet plasmid -> ran on gel | ||
+ | #*EncY looked fine, but Tet did not show up on gel | ||
+ | #Started overnight culture of Tet | ||
+ | |||
+ | =October 4, 2009= | ||
+ | #Miniprepped, digested Tet and ran on gel | ||
+ | #*Tet was confirmed | ||
+ | #gel extracted EncY, Tet and got low yields | ||
+ | #Started overnight ligations | ||
+ | #*1: Digested Tet + EncY + 5 | ||
+ | #*2: Gel extraced Tet + Digested EncY + 5 | ||
+ | #*negative controls were Tet plasmid alone | ||
+ | |||
+ | =October 8, 2009= | ||
+ | |||
+ | =October 9, 2009= | ||
+ | #Miniprepped EncY overnight and digested with E,P | ||
+ | #Ran digest on a gel | ||
+ | #*Enc part confirmed | ||
+ | #Transfected Enc+5 ligations from Sunday/Monday into competent cells from Invitrogen | ||
+ | #*Used NEB Hight Efficiency Transformation Protocol C2987 | ||
+ | #**did one 10-fold dilution | ||
+ | #**transformed 5ul of DNA | ||
+ | #**spread 100ul of cells on plate | ||
+ | #**shaker/incubator speed 232rpm | ||
+ | |||
+ | =October 11, 2009= | ||
+ | #Checked plates, no colonies yet | ||
+ | #Picked colonies and started overnights from an old 12E plate that wasn't checked: 12E i,ii,iii | ||
+ | |||
+ | =October 13, 2009= | ||
+ | #12E i and iii showed growth -> miniprepped | ||
+ | #Digested and ran on 1% agarose gel | ||
+ | #*Stained gel for 1 hour in EtBr+buffer | ||
+ | #*A faint ~700 band was observed! This could possibly be the 1+2+Enc insert that we want. | ||
+ | #To confirm the insert, we decided to sequence it | ||
+ | |||
+ | =October 15, 2009= | ||
+ | #Primers for sequencing 12E insert was designed and ordered | ||
+ | |||
+ | =October 16, 2009= | ||
+ | #EncY prepped and ready to ship down to MIT to submit part to registry |
Revision as of 20:23, 19 October 2009
Home | The Team | The Project | Parts Submitted to the Registry | Modeling | Bioinformatics | Notebook |
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Contents |
October 1, 2009
- Miniprepped Tet, 5, EncY, C
- Ran minipreps and K plasmid on a gel
- Besides Tet, all samples looked fine
- Poured K plates
October 2, 2009
- Digested K, EncY and ran on gel
October 3, 2009
- Digested EncY and old Tet plasmid -> ran on gel
- EncY looked fine, but Tet did not show up on gel
- Started overnight culture of Tet
October 4, 2009
- Miniprepped, digested Tet and ran on gel
- Tet was confirmed
- gel extracted EncY, Tet and got low yields
- Started overnight ligations
- 1: Digested Tet + EncY + 5
- 2: Gel extraced Tet + Digested EncY + 5
- negative controls were Tet plasmid alone
October 8, 2009
October 9, 2009
- Miniprepped EncY overnight and digested with E,P
- Ran digest on a gel
- Enc part confirmed
- Transfected Enc+5 ligations from Sunday/Monday into competent cells from Invitrogen
- Used NEB Hight Efficiency Transformation Protocol C2987
- did one 10-fold dilution
- transformed 5ul of DNA
- spread 100ul of cells on plate
- shaker/incubator speed 232rpm
- Used NEB Hight Efficiency Transformation Protocol C2987
October 11, 2009
- Checked plates, no colonies yet
- Picked colonies and started overnights from an old 12E plate that wasn't checked: 12E i,ii,iii
October 13, 2009
- 12E i and iii showed growth -> miniprepped
- Digested and ran on 1% agarose gel
- Stained gel for 1 hour in EtBr+buffer
- A faint ~700 band was observed! This could possibly be the 1+2+Enc insert that we want.
- To confirm the insert, we decided to sequence it
October 15, 2009
- Primers for sequencing 12E insert was designed and ordered
October 16, 2009
- EncY prepped and ready to ship down to MIT to submit part to registry