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Team:Cornell/Notebook/Salicylate reporter
From 2012.igem.org
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====June 29th, Friday==== | ====June 29th, Friday==== | ||
* [[Vent PCR]] at 11:00 (DPW) | * [[Vent PCR]] at 11:00 (DPW) | ||
| + | ** Amplifying both previous Phusion PCR band and original p21 template | ||
** Dylan's magic triple anneal method (55/60/63) | ** Dylan's magic triple anneal method (55/60/63) | ||
| - | * Gel purified PCR product (~1:20 pm) (DPW) | + | * Gel purified PCR product from Phusion template (~1:20 pm) (DPW) |
** Quantified product at 22.4 ng/uL | ** Quantified product at 22.4 ng/uL | ||
** Set up digestion of p21 PCR product with EcoRI-HF and AscI (~9:00 pm). | ** Set up digestion of p21 PCR product with EcoRI-HF and AscI (~9:00 pm). | ||
| Line 16: | Line 17: | ||
** Ran digestion on gel. (~11:00 pm) | ** Ran digestion on gel. (~11:00 pm) | ||
** Sliced out relevant band on gel, stored overnight at -20. | ** Sliced out relevant band on gel, stored overnight at -20. | ||
| - | * [[Miniprep]]ped overnight cultures of PL14-PL20 (STC) | + | |
| + | * [[Miniprep]]ped overnight cultures of PL14-PL20 (~1:00 pm, STC) | ||
** Using C1015 rotor, 6666 x g, the Corning culture tubes only fit in the centrifuge with the lids off | ** Using C1015 rotor, 6666 x g, the Corning culture tubes only fit in the centrifuge with the lids off | ||
| - | * Made [[LB]], 3x 60 mL in 100 mL bottles (SS) | + | |
| - | * Made [[LB|LB Agar]], 4 x 250 mL LB Agar in 500 mL bottles (CS) | + | * Made [[LB]], 3x 60 mL in 100 mL bottles (~ 3:00pm, SS) |
| - | * [[Autoclave]]d LB, LB Agar, and milliQ (SS) | + | * Made [[LB|LB Agar]], 4 x 250 mL LB Agar in 500 mL bottles (~3:00, CS) |
| - | * Made [[LB]] plates with Kan (CMR) | + | * [[Autoclave]]d LB, LB Agar, and milliQ (~3:30 pm, SS) |
| + | * Made [[LB]] plates with Kan (~6:30 pm, CMR) | ||
| + | |||
* CUGEM movie outing at 8:00 pm. | * CUGEM movie outing at 8:00 pm. | ||
| + | |||
* [[Phusion PCR]] at 10:00 pm (DPW) | * [[Phusion PCR]] at 10:00 pm (DPW) | ||
** Dylan's magic triple anneal method (57/65/70, 35 cycles total) | ** Dylan's magic triple anneal method (57/65/70, 35 cycles total) | ||
Revision as of 15:06, 30 June 2012
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June
Week
June 29th, Friday
- Vent PCR at 11:00 (DPW)
- Amplifying both previous Phusion PCR band and original p21 template
- Dylan's magic triple anneal method (55/60/63)
- Gel purified PCR product from Phusion template (~1:20 pm) (DPW)
- Quantified product at 22.4 ng/uL
- Set up digestion of p21 PCR product with EcoRI-HF and AscI (~9:00 pm).
- 22 ng/uL --> 45.5 uL sample for 1 ug digest
- Buffer 4
- Ran digestion on gel. (~11:00 pm)
- Sliced out relevant band on gel, stored overnight at -20.
- Miniprepped overnight cultures of PL14-PL20 (~1:00 pm, STC)
- Using C1015 rotor, 6666 x g, the Corning culture tubes only fit in the centrifuge with the lids off
- Made LB, 3x 60 mL in 100 mL bottles (~ 3:00pm, SS)
- Made LB Agar, 4 x 250 mL LB Agar in 500 mL bottles (~3:00, CS)
- Autoclaved LB, LB Agar, and milliQ (~3:30 pm, SS)
- Made LB plates with Kan (~6:30 pm, CMR)
- CUGEM movie outing at 8:00 pm.
- Phusion PCR at 10:00 pm (DPW)
- Dylan's magic triple anneal method (57/65/70, 35 cycles total)
- Amplifying nah operon from Gibson 1
- Appending BioBrick cutsites for ligation into pSB3C5
June 30th, Saturday
- Took PCR out of thermal cycler at 9:00 am (DPW)
- Set of gel using NEB 100bp and 2-log ladders (10:00am)
- Continued gel extraction of p21 PCR digest from previous day (SS)
- Set up ligation of p21 PCR digest extraction and dephosphorylated pBBRBB+mtrB (SS)
