Team:Freiburg/Notebook/23 August

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<a href="https://2011.igem.org/Team:Freiburg/Notebook/22_August">Previous entry</a>
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<a href="https://2011.igem.org/Team:Freiburg/Notebook"> 23 August </a>
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<a href="https://2011.igem.org/Team:Freiburg/Notebook/24_August">Next entry</a>
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==<span style="color:green;">green light receptor</span>==
==<span style="color:green;">green light receptor</span>==
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For a 50 µl reaction use
For a 50 µl reaction use
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P77: <br/>
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* CACAGGAAACCtcactaATGAGgATcCTTTTAGTGGAGGATGATTTGCc <br/>
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P78: <br/>
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* gGCAAATCATCCTCCACTAAAAGgATcCTCATtagtgaGGTTTCCTGTG <br/>
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<br/>
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===continuing with quickchanged CcaS and CcaR===
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'''Investigator: Julia'''
<br/>
<br/>
 +
on the plates for CcaR were no colonies at all :(<br/>
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the quickchanged CcaS had about six colonies per plate. <br/> nine were picked for miniprep ( 3ml LB medium with Kanamycin).<br/>
==<span style="color:blue;">blue light receptor</span>==
==<span style="color:blue;">blue light receptor</span>==
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'''Investigators: Sophie:'''
'''Investigators: Sophie:'''
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LOV-pcr, NOT-pcr and the Amp vector where ligated at 16°C
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LOV-pcr, NOT-pcr and the Amp vector where ligated for 1h at room temperature
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===Ligation===
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-
 
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'''Investigators: Sophie''' I ligated the purified inserts to the vector in the ratio 5:2 and 10:5 because of the loss of DNA due to the gelextraction
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===Transformation===
===Transformation===
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For a 50 µl reaction use
For a 50 µl reaction use
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P58:
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* TTCGAATTCGCGGCCGCTTCTAGATGGCCACCACCGTACAA <br/>
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P59:
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* CCGCTACTAGTATTATTACCCTTCTTTTGTCATGCCCT <br/>
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==<span style="color:grey;">Precipitator</span>==
==<span style="color:grey;">Precipitator</span>==
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===NAME OF YOUR EXPERIMENT===
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===Ligation===
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'''Investigators: Sophie'''
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Ligation of IPTG-promoter BBa_J04500, GFP-pbd and psb1c3-vector
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===Transformation===
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'''Investigators: NAME'''
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'''Investigators: Sophie'''
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Transformation of ligation product
{{:Team:Freiburg/Templates/footer}}
{{:Team:Freiburg/Templates/footer}}

Latest revision as of 01:11, 22 September 2011


This is the wiki page
of the Freiburger student
team competing for iGEM 2011.
Thank you for your interest!


Contents

green light receptor

PCR

Investigator: Jakob


PCR


Name:

Jakob

Date:

16.08.2011

Continue from Experiment: 12.08.2011

Order primers

Project Name:

Quickchange of CcaR (EcoRI)

PCR-Mixture for one Reaction:

For a 50 µl reaction use

P77:

  • CACAGGAAACCtcactaATGAGgATcCTTTTAGTGGAGGATGATTTGCc

P78:

  • gGCAAATCATCCTCCACTAAAAGgATcCTCATtagtgaGGTTTCCTGTG


32,5µl H20 Name
10µl 5x Phusion Buffer of Primer
2.5µl Primer fw P 77 (1:10)
2.5µl Primer dw P 78 (1:10)
1µl dNTPs of Template DNA
1µl DNA-Template CcaR from synechocystis 5ng/µl
0.5 µl Phusion (add in the end)

The program we used: for Quickchange of CcaR (EcoRI)


Temp.
Time
Nr. of cycles
94°
5’
1x
94°
30’’
2x
64°
30’’
72°
1’30’’
94°
30’’
2x
62°
30’’
72°
1’30’’
94°
30’’
5x
61,6°
30’’
72°
1’30’’
94°
30’’
25x
57,8°
30’’
72°
1’30’’
72°
7’
1x
4°
∞


continuing with quickchanged CcaS and CcaR

Investigator: Julia


on the plates for CcaR were no colonies at all :(
the quickchanged CcaS had about six colonies per plate.
nine were picked for miniprep ( 3ml LB medium with Kanamycin).

blue light receptor

Gel purification

Investigators:Sophie the bands of LOV-pcr and NOT-pcr where purificated. The vector was digested again because the band on the gel was not intense enough.

Ligation

Investigators: Sophie: LOV-pcr, NOT-pcr and the Amp vector where ligated for 1h at room temperature

Transformation

Investigators: Sophie: Transformation with the ligation product

red light receptor

PCR

Investigator: Jakob


PCR


Name:

Jakob

Date:

16.08.2011

Continue from Experiment: 12.08.2011

Order primers

Project Name:

Red light receptor, amplify Cph8

PCR-Mixture for one Reaction:

For a 50 µl reaction use

P58:

  • TTCGAATTCGCGGCCGCTTCTAGATGGCCACCACCGTACAA

P59:

  • CCGCTACTAGTATTATTACCCTTCTTTTGTCATGCCCT


32,5µl H20 Name
10µl 5x Phusion Buffer of Primer
2.5µl Primer fw P 58 (1:10)
2.5µl Primer dw P 59 (1:10)
1µl dNTPs of Template DNA
1µl DNA-Template JT122 5ng/µl
0.5 µl Phusion (add in the end)

The program we used: for Cph8



Temp.
Time
Nr. of cycles
94°
5’
1x
94°
30’’
2x
68°
30’’
72°
2’30’’
94°
30’’
2x
67°
30’’
72°
2’30’’
94°
30’’
5x
66°
30’’
72°
2’30’’
94°
30’’
25x
65°
30’’
72°
2’30’’
72°
7’
1x
4°
∞

Lysis cassette

NAME OF YOUR EXPERIMENT

Investigators:NAME


Precipitator

Ligation

Investigators: Sophie

Ligation of IPTG-promoter BBa_J04500, GFP-pbd and psb1c3-vector

Transformation

Investigators: Sophie Transformation of ligation product

                       

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