Team:UTP-Panama/After Regional Week 1

From 2011.igem.org

(Difference between revisions)
(→Octuber 17 to 23)
(→October 19th)
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<br>- auto-nail the eppendorf of 2 mL, they were ready.
<br>- auto-nail the eppendorf of 2 mL, they were ready.
<br>- prepare the Minimal Media for the experiments.
<br>- prepare the Minimal Media for the experiments.
 +
<br><br>
 +
For the new experiments:
 +
<br>- we used 5 mL of sustrates (liquid LB and MME)
 +
<br>- We did controls with and wothout antibiotics
 +
<br>- We used the following colonies:
 +
<br>  - JM109 (just the strain without plasmids)
 +
<br>  - with RFP
 +
<br>  - Bristol
 +
<br>  - GaTech
 +
<br>  - Renbo
 +
<br>- We did controls using the two methods:
 +
<br>  - Direct, studying the temperature
 +
<br>  - passing through the thermal shock, first.
 +
<br>- Thermic shock was:
 +
<br>  - @10ºC per 1 hour
 +
<br>-  Temperatures we used are:
 +
<br>  - 37ºC
 +
<br>  - between 15ºC and 20ºC  (18ºC)
 +
<br>- We needed 5 days of experimentation.
 +
<br><br> for each temperature and BioBrick we wanted to get (en each experiment):
 +
<br>  - growing curve
 +
<br>  - measurement of GFP expression
 +
<br>  - measurement of RFP expression, as control
 +
<br>  - measurement of heat production (thermometer)
 +
 +
<div align="center">Experiments:
 +
<br>
 +
<br>#1: 5 colonies in LB and MM in blank, for references without antibiotics and without nitrate, @37ºC and @18ºC.
 +
<br>#2: 5 colonies in LB and MM with nitrate @37ºC and @18ºC.
 +
<br>#3: 5 colonies in LB and MM with antibiotics @37ºC and @18ºC.
 +
<br>#4: 5 colonies in LB and MM with antibiotics and nitrate @18ºC, first passing through thermal shock and @18ºC without thermal shock, direct growing.
 +
<br>#5: 5 colonies in LB and MM with antibiotics and nitrate @37ºC, first passing through thermal shock and @37ºC without thermal shock, direct growing.
 +
<br><br>
 +
For Thursday, October 20th,  2011:
 +
<br>- Transforms competent cells with:
 +
<br>  - Bristol
 +
<br>  - Renbo, ligated with ampicillin
 +
<br><br>
 +
For Friday, October 21st, 2011:
 +
<br>  - take the plates out from the incubator
 +
<br>  - grow a Renbo colony, in liquid LB @37ºC with shaker.
 +
<br><br>
 +
For Saturday, October 22nd, 2011:
 +
<br>  - miniprep. of Renbo
 +
<br>  - electrophoresis of Renbo.

Revision as of 00:16, 29 October 2011

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October 17 to 23

October 18th

INVENTORY:

Quantity Description
24 petri plates (60 x 15 mm)
a lot! eppendorf tubes of 0,5 mL
4 tubes with linearized plasmids (kit)
8 tubes with competent cells
- genetic material from minipreparations
? ligation kit
2 tubes with SOC media (15 mL)
2 tubes with cloramphenicol (liquid or powder)
2 petri dishes with GaTech colonies
1 petri plates with RFP colonies
1 jar MgCl2 (for doing comp. cells)
1 jar CaCl2 (for doing comp. cells)
1 jar with ultrapure water
3 jars minipreparation Buffers (P1,P2,P3)
3 boxes small pipette tips (10 uL)
4 boxes medium pipette tips (200 uL)
3 boxes large pipette tips (1000 uL)
35 falcon tubes of 15 mL

Missing:
- large gloves
- falcon tubes 50 mL
- antibiotics
- LB media
- Eppendorf 2 mL for auto-nailing

October 19th

Ricardo reviewed the inventory and provided us:
- 1 box of large gloves
- 1 bag with falcons 50 mL
- 1 bag with eppendorf of 2 mL

Also:
- solid LB works
- not necessary to do liquid LB (there is)
- we have to transform competent cells with:
- Bristol (the plate isn't so good)
- RENBO (the product was ligated in the ampicillin plasmid) --> to do minipreparation and electrophoresis
- Colonies only without plasmids.

Plates with RFP and GaTech are fine!

We had to:
- prepare new antibiotics
- auto-nail the eppendorf of 2 mL, they were ready.
- prepare the Minimal Media for the experiments.

For the new experiments:
- we used 5 mL of sustrates (liquid LB and MME)
- We did controls with and wothout antibiotics
- We used the following colonies:
- JM109 (just the strain without plasmids)
- with RFP
- Bristol
- GaTech
- Renbo
- We did controls using the two methods:
- Direct, studying the temperature
- passing through the thermal shock, first.
- Thermic shock was:
- @10ºC per 1 hour
- Temperatures we used are:
- 37ºC
- between 15ºC and 20ºC (18ºC)
- We needed 5 days of experimentation.

for each temperature and BioBrick we wanted to get (en each experiment):
- growing curve
- measurement of GFP expression
- measurement of RFP expression, as control
- measurement of heat production (thermometer)

Experiments:



#1: 5 colonies in LB and MM in blank, for references without antibiotics and without nitrate, @37ºC and @18ºC.
#2: 5 colonies in LB and MM with nitrate @37ºC and @18ºC.
#3: 5 colonies in LB and MM with antibiotics @37ºC and @18ºC.
#4: 5 colonies in LB and MM with antibiotics and nitrate @18ºC, first passing through thermal shock and @18ºC without thermal shock, direct growing.
#5: 5 colonies in LB and MM with antibiotics and nitrate @37ºC, first passing through thermal shock and @37ºC without thermal shock, direct growing.

For Thursday, October 20th, 2011:
- Transforms competent cells with:
- Bristol
- Renbo, ligated with ampicillin

For Friday, October 21st, 2011:
- take the plates out from the incubator
- grow a Renbo colony, in liquid LB @37ºC with shaker.

For Saturday, October 22nd, 2011:
- miniprep. of Renbo


- electrophoresis of Renbo.
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