Team:Newcastle/24 August 2010

From 2010.igem.org

(Difference between revisions)
(Materials and Protocol)
(Single digestion of pSB1C3)
Line 67: Line 67:
Gel electrophoresis will be done on the 25th August, 2010.
Gel electrophoresis will be done on the 25th August, 2010.
-
 
-
=Single digestion of pSB1C3=
 
-
 
-
==Aims==
 
-
 
-
To linearise the vector pSB1C3. The linear vector will be used as a PCR template for Gibson cloning of the Subtilin immunity and ''rocF'' BioBricks.
 
-
 
-
==Materials and Protocol==
 
-
 
-
We are doing a single digest with HinDIII.
 
-
 
-
Please refer to [[Team:Newcastle/Restriction_digests|restriction digests]] and [[Team:Newcastle/Gel_electrophoresis|gel electrophoresis]].
 
-
 
-
==Results==
 
-
 
-
Gel electrophoresis results for digestion:
 
-
 
-
[[Image:24.08.10.png|500px]]
 
-
 
-
'''Figure 1''': Gel electrophoresis of the amplified PCR products and restriction digest
 
-
 
-
*'''Lane 1''': 1 Kb ladder
 
-
*'''Lane 2''': digested (linearised) pSB1C3
 
-
 
-
==Discussion and Conclusion==
 
-
 
-
The digestion was successful — the correct band was seen. We can proceed to gel extraction.
 
-
 
-
{{Team:Newcastle/footer}}
 

Revision as of 00:49, 28 October 2010

iGEM Homepage Newcastle University BacillaFilla Homepage Image Map

Contents

Miniprep for pGFPrrnB with filamentous cell part

Aims

The aim of this experiment is to obtain stock of the filamentous cell part in pGFP-rrnB for transformation into Bacillus subtilis 168.

Materials and Protocol

Please refer to qiagen minipreps and nanodrop spectrophotometer protocols.

Results

Tube 1 Tube 2 Tube 3 Tube 4 Tube 5 Tube 6 Tube 7 Tube 8 Tube 9 Tube 10 Tube 11 Tube 12
286.1 µl/ml 304.0 µl/ml 316.3 µl/ml 421.6 µl/ml 518.7 µl/ml 460.1 µl/ml 370.1 µl/ml 377.3 µl/ml 346.0 µl/ml 347.4 µl/ml 202.8 µl/ml 307.4 µl/ml

Table 1: Nanodrop spectrophotometer results. Table represents the amount of plasmid present in µl/ml quantity.

Discussion

The concentration ranged from 286.1 µl/ml to 518.7 µl/ml. Therefore we have obtained high concentration of yneA in pGFPrrnB.

Single and Double Digestion of pGFPrrnB with yneA

Aims

The aim of this experiment is to check if the insert yneA has been inserted into vector pGFPrrnB.

Materials and Protocol

We are doing two digests for pGFPrrnB and yneA:

  • Single digest with HindIII;
  • Double digest with EcoRI and NheI.


Please refer to restriction digests and gel electrophoresis.

Discussion

Gel electrophoresis will be done on the 25th August, 2010.