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Team:Cornell/Week 9
From 2011.igem.org
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(→Thursday, August 4) |
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:'''Objective''' | :'''Objective''' | ||
| - | ::Ligation of GFP | + | ::Ligation of GFP, RFP, and VioE genes with Avi-Tagged pZE12 backbone |
:'''Ligation''' | :'''Ligation''' | ||
| - | ::*''GFP + Avi- | + | ::*''GFP + Avi-Tagged Backbone'' |
| - | ::::10.6μL pZE12 | + | ::::10.6μL Avi-Tagged pZE12 vector backbone |
::::3.6μL GFP | ::::3.6μL GFP | ||
::::2.9μL H2O | ::::2.9μL H2O | ||
| Line 28: | Line 28: | ||
::::<u>1.0μL T4 DNA Ligase</u> | ::::<u>1.0μL T4 DNA Ligase</u> | ||
::::20μL Total | ::::20μL Total | ||
| - | ::*''RFP + Avi- | + | ::*''RFP + Avi-Tagged Backbone'' |
| - | ::::10.3μL pZE12 | + | ::::10.3μL Avi-Tagged pZE12 vector backbone |
::::6.7μL RFP | ::::6.7μL RFP | ||
::::2.0μL T4 DNA Ligase Buffer | ::::2.0μL T4 DNA Ligase Buffer | ||
::::<u>1.0μL T4 DNA Ligase</u> | ::::<u>1.0μL T4 DNA Ligase</u> | ||
::::20μL Total | ::::20μL Total | ||
| - | ::*''VioE + Avi- | + | ::*''VioE + Avi-Tagged Backbone'' |
::::2.3μL H2O | ::::2.3μL H2O | ||
::::4.1μL VioE | ::::4.1μL VioE | ||
| - | ::::10.6μL pZE12 | + | ::::10.6μL Avi-Tagged pZE12 vector backbone |
::::2.0μL T4 DNA Ligase Buffer | ::::2.0μL T4 DNA Ligase Buffer | ||
::::<u>1.0μL T4 DNA Ligase</u> | ::::<u>1.0μL T4 DNA Ligase</u> | ||
| Line 44: | Line 44: | ||
::Prepare control ligation reactions (inserts replaced with H2O) for each of the above constructs. | ::Prepare control ligation reactions (inserts replaced with H2O) for each of the above constructs. | ||
:::*Same volume of vector backbone, buffer, ligase | :::*Same volume of vector backbone, buffer, ligase | ||
| - | :::*Volumes of all inserts (i.e. gene | + | :::*Volumes of all inserts (i.e. gene) go toward volume of H2O |
::After ligation setup, incubate in 16°C waterbath overnight. | ::After ligation setup, incubate in 16°C waterbath overnight. | ||
Revision as of 21:25, 4 August 2011
Week 1 | Week 2 - 5 | Week 6 | Week 7 | Week 8 | Week 9 | Week 10 | Week 11 | Week 12 | Week 13 | Week 14 | Week 15 | Week 16 | Week 17 | Week 18 | Week 19 | Week 20 | Week 21 |
July 31st - August 6th
Sunday
Monday
Tuesday
Wednesday
Thursday, August 4
Afternoon lab work done by: Claire Paduano, Charlie Chung
- Objective
- Ligation of GFP, RFP, and VioE genes with Avi-Tagged pZE12 backbone
- Ligation
- GFP + Avi-Tagged Backbone
- 10.6μL Avi-Tagged pZE12 vector backbone
- 3.6μL GFP
- 2.9μL H2O
- 2.0μL T4 DNA Ligase Buffer
- 1.0μL T4 DNA Ligase
- 20μL Total
- RFP + Avi-Tagged Backbone
- 10.3μL Avi-Tagged pZE12 vector backbone
- 6.7μL RFP
- 2.0μL T4 DNA Ligase Buffer
- 1.0μL T4 DNA Ligase
- 20μL Total
- VioE + Avi-Tagged Backbone
- 2.3μL H2O
- 4.1μL VioE
- 10.6μL Avi-Tagged pZE12 vector backbone
- 2.0μL T4 DNA Ligase Buffer
- 1.0μL T4 DNA Ligase
- 20μL Total
- In all above reactions, volume of plasmid vector added corresponds with 100ng backbone.
- Prepare control ligation reactions (inserts replaced with H2O) for each of the above constructs.
- Same volume of vector backbone, buffer, ligase
- Volumes of all inserts (i.e. gene) go toward volume of H2O
- After ligation setup, incubate in 16°C waterbath overnight.
