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Team:Newcastle/Labwork/25 September 2009
From 2009.igem.org
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Formal Lab Session - 25th September 2009
Chassis team
Introduction
- To obtain pspac part from pMutin4, we digested psc fragment with HindIII and BamHI
- We need Mini prep kinA transformation and check the result.
Experiment procedure
Digestion
- Digest psc clean-up DNA
dd H2O 7ul
10X fast digest buffer 7ul
Fast HindIII 3ul
Fast BamHI 3ul
psc clean-up DNA 50ul
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70ul
- Incubate 1 hour at 37 degree
Gel Extraction
- After the digestion of psc fragment we run the digested DNA on big well gel and cut the right band and purified the DNA by gel extraction kit.
Mini prep
- Followed the Phil's protocal.
- Since we cultured 24 colonies yesterday, we performed Mini Prep for all those 24 strains.
Test the Mini Prep result
- Digested the Mini Prep DNA with EcoRI and NheI
dd H2O 7ul
10X fast digest buffer 2ul
Fast EcoRI 0.5ul
Fast NheI 0.5ul
Mini Prep DNA 10ul
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20ul
- 37C incubated for 1 hour
- Add 2ul stop buffer and run the sample on agarose gel.
Ligation
- Ligate psc fragment with pMutin4 backbone
ddH2O 10ul
ligation buffer 4ul
psc fragments 10ul
pMutin4 backbone (24.9ng/ul) 5ul
T4 ligase 1ul
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30ul
conclusion
- After the digestion of kinA transformation Mini Prep result, it seems No.4,5,6,10 could be the right clone. So, we prepare Midi culture for all 4 strains.
Futher plan
- Use HindIII enzyme digest Mini prep result of those 4 strains to double check the result.
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News
Events
- 20 – 21 June 2009 - Europe workshop (London)
- 23 – 24 June 2009 - UK iGEM meetup (Edinburgh)
- 23 October Practice Presentation (Newcastle)
- 23 October T-shirts are ready
- 27 October Practice Presentation (Sunderland)
- 27 October Poster is ready
- 30 October – 2 November 2009 - Jamboree (Boston)
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