Team:Warsaw/Calendar-Main/28 August 2009

Revision as of 20:48, 30 August 2009 by Messer (Talk | contribs)

Tasks:

Methods:

The PCR mix was prepared as follows:

5μl buffer B (EURx)
2μl 5mM dNTPs (EURx)
5μl forward starter
5μl reverse starter
2μl OptiTaq polymerase (EURx)
 2μl Salmonella matrix
 29 μl H₂O

PCR programme:

 4min 95°C 
(30s 95°C, 35s 48°C, 40s 72°C)x28
 (30s 95°C, 35s 58°C, 40s 72°C)x28 
 10min 72°C 
 ~ 4°C

The digest mix was prepared as follows:

60μl plasmid isolation
7μl Buffer O (Fermentas)
1μl EcoRI enzyme
1μl PstI enzyme
1μl H₂O

The digest was carried out in 37°C for 3h and inactivated for 15 min. in 80°C.
1μl of CIAP enzyme was added after the inactivation and allowed to work for 1h in 37°C and was later inactivated for 15 min. in 85°C.
The resulting mix was separated on 1% agarose gel alongside the PCR and then selected bands were extracted for purification.

Results:

No gel picture, but the PCR worked so great that I decided not to irradiate the gel with UV light more than it was necessary for band extraction.

Conclusions:

Tasks:

Methods:

The digest mix was prepared as follows:

20μl plasmid isolation
3μl Buffer O (Fermentas)
1μl EcoRI enzyme
1μl PstI enzyme
5μl H₂O