Team:Paris Saclay/Notebook/August/20

From 2014.igem.org

(Difference between revisions)
(Bacterial culture)
(CAD PCR)
Line 47: Line 47:
Very small quantity of DNA
Very small quantity of DNA
-
1.5 µl --> Gotaq
+
1.5 µl --> DMSO
-
0.25
+
0.25 µl --> Gotaq
 +
 
 +
PCR cycle
 +
 
 +
95°  2 min
 +
95°  30 sec
 +
58°  30 sec
 +
72°  1 min 45
 +
72°  5 min
 +
 
 +
 
 +
 
 +
====B - Construction of the fusion protein (color)=====
 +
PCR with Gotaq
 +
 
 +
22.75 µl --> H2O
 +
 
 +
10 µl --> 5xbuffer
 +
 
 +
1 µl --> dNTP
 +
 
 +
4 µl --> MgCl2
 +
 
 +
5 µl iPS 83 (Primer)
 +
 
 +
5 µl iPS 84
 +
 
 +
0.5 µl --> DNA
 +
 
 +
1.5 µl --> DMSO
 +
 
 +
0.25 µl --> Gotaq
 +
 
 +
 
 +
PCR cycle
 +
 
 +
95°  2 min
 +
95°  30 sec
 +
58°  30 sec
 +
72°  1 min 45
 +
72°  5 min

Revision as of 14:38, 20 August 2014

Contents

Wednesday 20th August

Back to the calendar

D- Lemon scent

Bacterial culture

by Laetitia

Here are the results of the bacterial culture of yesterday

20 08 bacterial culture.jpg

Bacteria with the pGEMTeasy without the DNA insert are blue and the bacteria with the pGEMTeasy and the DNA insert (PS, GS or LS gene) are white

Then, we have chosen 3 white colonies for each DNA insert and we have launched 3 corresponding liquid cultures :

- 5mL LB + ampiciline (1/1000e)

- 1 white colony

The 9 cultures are incubated at 37°C


by Melanie

Preculture 20ml of DH5a pPSI (apra 1/2000)


CAD PCR

Today we have receive our synthetic gene CAD (cinnamyl alcohol deshydrogenase) :)

so we do a PCR (with go taq):

23.25 µl --> H2O

10 µl --> 5xbuffer

1 µl --> dNTP

4 µl --> MgCl2

5 µl iPS 79 (Primer)

5 µl iPS 80

Very small quantity of DNA

1.5 µl --> DMSO

0.25 µl --> Gotaq

PCR cycle

95° 2 min 95° 30 sec 58° 30 sec 72° 1 min 45 72° 5 min


B - Construction of the fusion protein (color)=

PCR with Gotaq

22.75 µl --> H2O

10 µl --> 5xbuffer

1 µl --> dNTP

4 µl --> MgCl2

5 µl iPS 83 (Primer)

5 µl iPS 84

0.5 µl --> DNA

1.5 µl --> DMSO

0.25 µl --> Gotaq


PCR cycle

95° 2 min 95° 30 sec 58° 30 sec 72° 1 min 45 72° 5 min