Team:Paris/20 August 2009
From 2009.igem.org
(→Lab work) |
(→Lab work) |
||
(7 intermediate revisions not shown) | |||
Line 4: | Line 4: | ||
==NoteBook== | ==NoteBook== | ||
- | { | + | {{Paris2009_Calendar}} |
- | + | {{Paris2009_Calendar_Link|19_August_2009|21_August_2009}} | |
- | + | <center> '''August 20th''' </center> | |
- | + | ||
- | + | ||
- | + | ||
- | + | ||
<html> | <html> | ||
Line 18: | Line 14: | ||
<div id="paris_content"> | <div id="paris_content"> | ||
</html> | </html> | ||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
==Lab work== | ==Lab work== | ||
Line 44: | Line 25: | ||
<u>'''''Pcr of''''':</u> | <u>'''''Pcr of''''':</u> | ||
- | A35 - FecI from pSV26 | + | A35 - FecI from pSV26 '''Worked''' |
- | A35 bis- FecI from K12 genomic DNA source | + | A35 bis- FecI from K12 genomic DNA source '''Failed''' (Skywalker is only a padawan) |
</div> | </div> | ||
Line 67: | Line 48: | ||
</div> | </div> | ||
- | < | + | |
+ | <div class="vicard"> | ||
+ | Digestion | ||
</div> | </div> | ||
- | <div | + | <div class="experience"> |
+ | |||
+ | Digestion in XP | ||
+ | |||
+ | A30->D40 (Nter RFP) | ||
+ | |||
+ | A31->D41 (RFP) | ||
+ | |||
+ | A33->D42 (Cter RFP) | ||
+ | |||
+ | with: | ||
+ | -20µL DNA | ||
+ | -2µL Xba | ||
+ | -2µL PST | ||
+ | -0,5µL BSA*100 | ||
+ | -5µL Buffer 2*10 | ||
+ | -20,5µL H20 | ||
+ | |||
+ | Then purification | ||
</div> | </div> | ||
- | |||
- | |||
- | == | + | <div class="vicard"> |
+ | Ligation | ||
+ | </div> | ||
+ | <div class="experience"> | ||
+ | '''Ligation''' | ||
+ | '''DO:''' | ||
- | + | -PSB1A3: 0,61 µg/ml | |
+ | |||
+ | -D40:0,50 µg/ml | ||
+ | |||
+ | -D41:0,80 µg/ml | ||
+ | |||
+ | -D42:0,40 µg/ml | ||
+ | |||
+ | '''D40 with PSB1A3 in 3X and 10x''' | ||
+ | |||
+ | For 3x(3,7µl insert+1µL vector+1µL buffer 10*+1µl ligase+3,3µL H2O) | ||
+ | |||
+ | For 10x(12µl insert+1µL vector+2µL buffer 10*+1µl ligase+4µL H2O) | ||
+ | |||
+ | '''D41 with PSB1A3 in 3x and 10x''' | ||
+ | |||
+ | For 3x(2,5µl insert+1µL vector+1µL buffer 10*+1µl ligase+4,5µL H2O) | ||
+ | |||
+ | For 10x(7,5µl insert+1µL vector+1,5µL buffer 10*+1µl ligase+4µL H2O) | ||
+ | |||
+ | '''D42 with PSB1A3 in 3x and 10x''' | ||
+ | |||
+ | For 3x(3,7µl insert+1µL vector+1µL buffer 10*+1µl ligase+3,3µL H2O) | ||
+ | |||
+ | For 10x(12µl insert+1µL vector+2µL buffer 10*+1µl ligase+4µL H2O) | ||
+ | |||
+ | '''For negativ control:''' | ||
+ | |||
+ | 0µl insert+1µL vector+2µL buffer 10*+1µl ligase+7µL H2O | ||
+ | |||
+ | |||
+ | |||
+ | </div> | ||
+ | {{Paris2009_Calendar_Link|19_August_2009|21_August_2009}} |
Latest revision as of 22:36, 16 October 2009
NoteBook
|
|
|
|
|
---|
Lab work
PCR w/ Skywalker
Pcr of:
A35 - FecI from pSV26 Worked
A35 bis- FecI from K12 genomic DNA source Failed (Skywalker is only a padawan)
Purification of Pcr product
directly after PCR:
IMAGE
After purification :
IMAGE (wait until tomorrow)
Digestion
Digestion in XP
A30->D40 (Nter RFP)
A31->D41 (RFP)
A33->D42 (Cter RFP)
with: -20µL DNA -2µL Xba -2µL PST -0,5µL BSA*100 -5µL Buffer 2*10 -20,5µL H20
Then purification
Ligation
Ligation DO:
-PSB1A3: 0,61 µg/ml
-D40:0,50 µg/ml
-D41:0,80 µg/ml
-D42:0,40 µg/ml
D40 with PSB1A3 in 3X and 10x
For 3x(3,7µl insert+1µL vector+1µL buffer 10*+1µl ligase+3,3µL H2O)
For 10x(12µl insert+1µL vector+2µL buffer 10*+1µl ligase+4µL H2O)
D41 with PSB1A3 in 3x and 10x
For 3x(2,5µl insert+1µL vector+1µL buffer 10*+1µl ligase+4,5µL H2O)
For 10x(7,5µl insert+1µL vector+1,5µL buffer 10*+1µl ligase+4µL H2O)
D42 with PSB1A3 in 3x and 10x
For 3x(3,7µl insert+1µL vector+1µL buffer 10*+1µl ligase+3,3µL H2O)
For 10x(12µl insert+1µL vector+2µL buffer 10*+1µl ligase+4µL H2O)
For negativ control:
0µl insert+1µL vector+2µL buffer 10*+1µl ligase+7µL H2O