Imperial College/23 September 2008

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(New page: ==Dry Lab== *Modelling Motility **Done up the code to model the cell's trajectory using least square curve fit method. **Fitted cell trajectories of 100908 Video 15. Results look promisin...)
 
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|{{#calendar: title=Imperial_College |year=2008 | month=08}}
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|{{#calendar: title=Imperial_College |year=2008 | month=09}}
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|{{#calendar: title=Imperial_College |year=2008 | month=10}}
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=23 September 2008=
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==Wet Lab==
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===Single Colony PCR===
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[[Image:PCR-23.PNG|center|500px]]
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*In order to verify the ligations of parts (from geneart and PCR products) into the biobrick vector AK3 we have carried out a series of single colony PCRs using the primers Psb.
 +
*The conditions used were as follows:
 +
**1 cycle - 95<sup>o</sup>C for 30 seconds
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**30 cycles - 95<sup>o</sup>C for 30 seconds, 60<sup>o</sup>C for 30 seconds,72<sup>o</sup>C for 30 seconds
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**1 cycle - 72<sup>o</sup>C for 2 minutes
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*The numbers of the ligations correspond to the following ligation reactions:
 +
**6 L  (from geneart construct 6) = LipA-Elastin
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**EpsE (from geneart construct 3) = EpsE gene
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**AmyE5' (from PCR products)
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**AmyE3' (from PCR products)
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**8<sub>1</sub> (from geneart constructs 6, PCR using mini DNA) =  pGsiB-gsiB
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**8<sub>2</sub> (from geneart constructs 6, PCR using mini DNA) = pGsiB-gsiB
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**Negative contains no DNA,
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*The results show positive results for EpsE, Amy5',AmyE3', 8 (pGsiB-gsiB)
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*The result from AmyE3' is less clear, it appears that the 1st of the three is the correct size but all will be checked by mini-preping and then digestion.
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*There was no contamination in the negative control.
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===Miniprep Digestion===
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*All minpreps from yesterday digested with ''Eco''RI and ''Pst''I
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[[Image:Digest23-9.png|center|500px]]
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*Each lane is a separate miniprep (2 minipreps of each transformation).
 +
*Band from the EpsE digests is approximately correctly, although this is also approxiamtely the size of GFP-Termiantor and RFP-Terminator. In particular, the pSB1AK3 vector containing RFP-T was used to provide vector for the ligation.
 +
*All other mini-preps do not appear to have inserts! The gel had been imaged earlier and had also shown no evidence of inserts.
 +
*It should also be noted that the vector in all the minipreps appears to be the wrong size!
 +
==Dry Lab==
==Dry Lab==
*Modelling Motility
*Modelling Motility
**Done up the code to model the cell's trajectory using least square curve fit method.  
**Done up the code to model the cell's trajectory using least square curve fit method.  
**Fitted cell trajectories of 100908 Video 15. Results look promising:
**Fitted cell trajectories of 100908 Video 15. Results look promising:
 +
[[Image:V15Cell1.JPG|thumb|500px|center|Video 15 Cell 1]]
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[[Image:V15Cell5.jpg|thumb|500px|center|Video 15 Cell 5]]
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<br>
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{{Imperial/EndPage|Notebook|Notebook}}

Latest revision as of 20:48, 28 October 2008

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23 September 2008

Wet Lab

Single Colony PCR

PCR-23.PNG
  • In order to verify the ligations of parts (from geneart and PCR products) into the biobrick vector AK3 we have carried out a series of single colony PCRs using the primers Psb.
  • The conditions used were as follows:
    • 1 cycle - 95oC for 30 seconds
    • 30 cycles - 95oC for 30 seconds, 60oC for 30 seconds,72oC for 30 seconds
    • 1 cycle - 72oC for 2 minutes
  • The numbers of the ligations correspond to the following ligation reactions:
    • 6 L (from geneart construct 6) = LipA-Elastin
    • EpsE (from geneart construct 3) = EpsE gene
    • AmyE5' (from PCR products)
    • AmyE3' (from PCR products)
    • 81 (from geneart constructs 6, PCR using mini DNA) = pGsiB-gsiB
    • 82 (from geneart constructs 6, PCR using mini DNA) = pGsiB-gsiB
    • Negative contains no DNA,
  • The results show positive results for EpsE, Amy5',AmyE3', 8 (pGsiB-gsiB)
  • The result from AmyE3' is less clear, it appears that the 1st of the three is the correct size but all will be checked by mini-preping and then digestion.
  • There was no contamination in the negative control.

Miniprep Digestion

  • All minpreps from yesterday digested with EcoRI and PstI
Digest23-9.png
  • Each lane is a separate miniprep (2 minipreps of each transformation).
  • Band from the EpsE digests is approximately correctly, although this is also approxiamtely the size of GFP-Termiantor and RFP-Terminator. In particular, the pSB1AK3 vector containing RFP-T was used to provide vector for the ligation.
  • All other mini-preps do not appear to have inserts! The gel had been imaged earlier and had also shown no evidence of inserts.
  • It should also be noted that the vector in all the minipreps appears to be the wrong size!

Dry Lab

  • Modelling Motility
    • Done up the code to model the cell's trajectory using least square curve fit method.
    • Fitted cell trajectories of 100908 Video 15. Results look promising:
Video 15 Cell 1
Video 15 Cell 5
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