Team:Stockholm/4 August 2010

From 2010.igem.org

(Difference between revisions)
(New page: {{Stockholm/Top2}} ==Andreas==)
(→Andreas)
Line 2: Line 2:
==Andreas==
==Andreas==
 +
 +
===Cloning results===
 +
''From 3/8 transformations''
 +
 +
{|border="1" cellpadding="2" cellspacing="0" width="300"
 +
|width="120"|'''pSB1A3.SOD:'''
 +
|rowspan="3"|Bacterial lawn on all three plates, probably due to the Amp in plates having degraded. Plates discarded
 +
|-
 +
|'''pSB1A3.yCCS A:'''
 +
|-
 +
|'''pSB1A3.yCCS B:'''
 +
|-
 +
|'''pSB1C3.IgG prot.:'''
 +
|Good colony yield. Left to grow and mature for 4-5 more hours until red colonies appeared.
 +
|}
 +
 +
===ON cultures===
 +
Four colonies picked from IgG protease plate and resuspended in 10 μl LB. 3 μl used to inoculate 5 ml LB + 25 Cm. Grown ON in 37 °C, 250 rpm.
 +
 +
===Cloning/transformations===
 +
New quick-transformations were performed for:
 +
*pSB1A3.SOD
 +
*pSB1A3.yCCS A
 +
*pSB1A3.yCCS B
 +
 +
Procedures as described 3/8.
 +
 +
===CPP DNA synthesis===
 +
[[Image:CPP_C_cluster.jpg|200px|thumb|right|Cluster of C-CPPs.]]
 +
[[Image:CPP_N_cluster.jpg|200px|thumb|right|Cluster of N-CPPs.]]
 +
Since our previous CPP synthesis order was rejected due to repetetive sequences Johan and I redesigned the order into two separate clusters, CPP_N and CPP_C.
 +
The new clusters were designed with both restriction sites and primer annealing sites flanking each CPP. An extension sequence was also added to enable gel separation after digestion, as the CPPs are very similar in length.
 +
 +
Johan also redesigned the coding sequence by shifting codons by the addition of "silent mutations".

Revision as of 17:18, 5 August 2010

Home Project Idea Lab Work Results Modelling Team       Follow us on and        2010.igem.org

↳   Notebook Safety Protocols


Contents

Andreas

Cloning results

From 3/8 transformations

pSB1A3.SOD: Bacterial lawn on all three plates, probably due to the Amp in plates having degraded. Plates discarded
pSB1A3.yCCS A:
pSB1A3.yCCS B:
pSB1C3.IgG prot.: Good colony yield. Left to grow and mature for 4-5 more hours until red colonies appeared.

ON cultures

Four colonies picked from IgG protease plate and resuspended in 10 μl LB. 3 μl used to inoculate 5 ml LB + 25 Cm. Grown ON in 37 °C, 250 rpm.

Cloning/transformations

New quick-transformations were performed for:

  • pSB1A3.SOD
  • pSB1A3.yCCS A
  • pSB1A3.yCCS B

Procedures as described 3/8.

CPP DNA synthesis

Cluster of C-CPPs.
Cluster of N-CPPs.

Since our previous CPP synthesis order was rejected due to repetetive sequences Johan and I redesigned the order into two separate clusters, CPP_N and CPP_C. The new clusters were designed with both restriction sites and primer annealing sites flanking each CPP. An extension sequence was also added to enable gel separation after digestion, as the CPPs are very similar in length.

Johan also redesigned the coding sequence by shifting codons by the addition of "silent mutations".

Retrieved from "http://2010.igem.org/Team:Stockholm/4_August_2010"