Team:Freiburg/Notebook/7 September

From 2011.igem.org

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==<span style="color:orange;">Lysis cassette</span>==
==<span style="color:orange;">Lysis cassette</span>==
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===NAME OF YOUR EXPERIMENT===
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===Troubleshooting of the modified Lysis genes K124017===
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'''Investigators:NAME'''
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'''Investigators:Theo'''
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<br>
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'''''S4+S15 Nr1 and 7''''' were digested with '''''XbaI''''' and '''''PstI''''', loaded on a 1% agarose gel and extracted on the 6th of September.
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<br>
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Today I digested the quickchanged-temperature sensitive promotor (K098995) with '''''SpeI''''' and '''''PstI''''' for ca 3 hours, incubated it with antarctic phosphatase for 1 hour and PCR-purified it.
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==<span style="color:grey;">Precipitator</span>==
==<span style="color:grey;">Precipitator</span>==

Revision as of 17:29, 7 September 2011


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

Contents

Commons

Ligation

Investigators: Sandra

Ligation of PR1-PR6 with RFP (S1b and S2a). PR1-PR6 were digested with antarctica before start of ligation. Start of ligation at 12:40. Incubation at room temperature for at least 3 hours.

Trafo

Investigators: Sandra

Trafo of:

  • PR1+GFP
  • PR2+GFP
  • PR3+GFP
  • PR4+GFP
  • PR5+GFP
  • PR6+GFP
  • PR1+50b
  • PR2+50b

green light receptor

NAME OF YOUR EXPERIMENT

Investigators:NAME


blue light receptor

no colonies on the plates

There were no colonies on the plates with ligated parts of the 2A-assembly.

Miniprep

Investigators: Sophie
yesterday two tubes with LB were inoculated with cells from our S35 glycerol stock(BBa_K322999). Today minipreps were performed.

PCR

Investigators: Sophie

Name: Sophie


Date: 07.09.11
Continue from Experiment (Date)

(Name)

Project Name: Gibson of ♥ and NOT

PCR-Mixture for one Reaction:

For a 50 µl reaction use


32,5µl H20 Name
10µl 5x Phusion Buffer of Primer
2.5µl Primer fw ♥: P97

NOT: P99

2.5µl Primer dw ♥: P98

NOT: P100

1µl dNTPs of Template DNA
1µl DNA-Template M45a,b,c (BBa_Q0400)

Miniprep of M35 (1x) and M35b (2x) (BBa_K322999)

0.5 µl Phusion (add in the end)

Digestion with Dpn I and purification with PCR purification kit What program do you use?

First 20 cycles touchdown 69°C -0.4/cycle

next 10 cycles touchdown 72°C -0.2 per cycle


PCR products were loaded onto a gel

red light receptor

NAME OF YOUR EXPERIMENT

Investigators:NAME



Lysis cassette

Troubleshooting of the modified Lysis genes K124017

Investigators:Theo


S4+S15 Nr1 and 7 were digested with XbaI and PstI, loaded on a 1% agarose gel and extracted on the 6th of September.

Today I digested the quickchanged-temperature sensitive promotor (K098995) with SpeI and PstI for ca 3 hours, incubated it with antarctic phosphatase for 1 hour and PCR-purified it.

Precipitator

Trafo

Investigators: Sandra

Trafo of the already ligated parts of the pbd + Gst-vector

Ligation

Investigators: Sophie

For pbd in iGEM C3-vector: again Ligation (see 31.08.11) because I don't find the last ligation product that produced positive clones. I have to redo the Trafo because the last positive clones seem to have mutated over the last generations, probably because the pbd is toxic.
labelled: L S54-ε5-C3 1:1 / 1:3
stored in "Ligationsreaktionen"-box

                       

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