Imperial College/23 September 2008

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==Wet Lab==
==Wet Lab==
===Single Colony PCR===
===Single Colony PCR===
 +
[[Image:PCR-23.PNG|center|500px]]
 +
*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
 +
**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
 +
**1 cycle - 72<sup>o</sup>C 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)
 +
**8<sub>1</sub> (from geneart constructs 6, PCR using mini DNA) =  pGsiB-gsiB
 +
**8<sub>2</sub> (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===
===Miniprep Digestion===
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[[Image:V15Cell1.JPG|thumb|500px|center|Video 15 Cell 1]]
[[Image:V15Cell1.JPG|thumb|500px|center|Video 15 Cell 1]]
[[Image:V15Cell5.jpg|thumb|500px|center|Video 15 Cell 5]]
[[Image:V15Cell5.jpg|thumb|500px|center|Video 15 Cell 5]]
 +
<br>
 +
{{Imperial/EndPage|Notebook|Notebook}}

Latest revision as of 20:48, 28 October 2008

August
MTWTFSS
        [http://2008.igem.org/Imperial_College/1_August_2008 1] [http://2008.igem.org/Imperial_College/2_August_2008 2] [http://2008.igem.org/wiki/index.php?title=Imperial_College/3_August_2008&action=edit 3]
[http://2008.igem.org/Imperial_College/4_August_2008 4] [http://2008.igem.org/Imperial_College/5_August_2008 5] [http://2008.igem.org/wiki/index.php?title=Imperial_College/6_August_2008&action=edit 6] [http://2008.igem.org/Imperial_College/7_August_2008 7] [http://2008.igem.org/Imperial_College/8_August_2008 8] [http://2008.igem.org/wiki/index.php?title=Imperial_College/9_August_2008&action=edit 9] [http://2008.igem.org/Imperial_College/10_August_2008 10]
[http://2008.igem.org/Imperial_College/11_August_2008 11] [http://2008.igem.org/Imperial_College/12_August_2008 12] [http://2008.igem.org/Imperial_College/13_August_2008 13] [http://2008.igem.org/Imperial_College/14_August_2008 14] [http://2008.igem.org/Imperial_College/15_August_2008 15] [http://2008.igem.org/Imperial_College/16_August_2008 16] [http://2008.igem.org/Imperial_College/17_August_2008 17]
[http://2008.igem.org/Imperial_College/18_August_2008 18] [http://2008.igem.org/Imperial_College/19_August_2008 19] [http://2008.igem.org/Imperial_College/20_August_2008 20] [http://2008.igem.org/Imperial_College/21_August_2008 21] [http://2008.igem.org/Imperial_College/22_August_2008 22] [http://2008.igem.org/wiki/index.php?title=Imperial_College/23_August_2008&action=edit 23] [http://2008.igem.org/Imperial_College/24_August_2008 24]
[http://2008.igem.org/Imperial_College/25_August_2008 25] [http://2008.igem.org/Imperial_College/26_August_2008 26] [http://2008.igem.org/Imperial_College/27_August_2008 27] [http://2008.igem.org/Imperial_College/28_August_2008 28] [http://2008.igem.org/Imperial_College/29_August_2008 29] [http://2008.igem.org/wiki/index.php?title=Imperial_College/30_August_2008&action=edit 30] [http://2008.igem.org/Imperial_College/31_August_2008 31]
September
MTWTFSS
[http://2008.igem.org/Imperial_College/1_September_2008 1] [http://2008.igem.org/Imperial_College/2_September_2008 2] [http://2008.igem.org/Imperial_College/3_September_2008 3] [http://2008.igem.org/Imperial_College/4_September_2008 4] [http://2008.igem.org/Imperial_College/5_September_2008 5] [http://2008.igem.org/wiki/index.php?title=Imperial_College/6_September_2008&action=edit 6] [http://2008.igem.org/Imperial_College/7_September_2008 7]
[http://2008.igem.org/Imperial_College/8_September_2008 8] [http://2008.igem.org/Imperial_College/9_September_2008 9] [http://2008.igem.org/Imperial_College/10_September_2008 10] [http://2008.igem.org/Imperial_College/11_September_2008 11] [http://2008.igem.org/Imperial_College/12_September_2008 12] [http://2008.igem.org/wiki/index.php?title=Imperial_College/13_September_2008&action=edit 13] [http://2008.igem.org/wiki/index.php?title=Imperial_College/14_September_2008&action=edit 14]
[http://2008.igem.org/Imperial_College/15_September_2008 15] [http://2008.igem.org/Imperial_College/16_September_2008 16] [http://2008.igem.org/Imperial_College/17_September_2008 17] [http://2008.igem.org/Imperial_College/18_September_2008 18] [http://2008.igem.org/Imperial_College/19_September_2008 19] [http://2008.igem.org/wiki/index.php?title=Imperial_College/20_September_2008&action=edit 20] [http://2008.igem.org/wiki/index.php?title=Imperial_College/21_September_2008&action=edit 21]
[http://2008.igem.org/Imperial_College/22_September_2008 22] [http://2008.igem.org/Imperial_College/23_September_2008 23] [http://2008.igem.org/Imperial_College/24_September_2008 24] [http://2008.igem.org/Imperial_College/25_September_2008 25] [http://2008.igem.org/Imperial_College/26_September_2008 26] [http://2008.igem.org/wiki/index.php?title=Imperial_College/27_September_2008&action=edit 27] [http://2008.igem.org/wiki/index.php?title=Imperial_College/28_September_2008&action=edit 28]
[http://2008.igem.org/Imperial_College/29_September_2008 29] [http://2008.igem.org/Imperial_College/30_September_2008 30]
October
MTWTFSS
    [http://2008.igem.org/Imperial_College/1_October_2008 1] [http://2008.igem.org/Imperial_College/2_October_2008 2] [http://2008.igem.org/wiki/index.php?title=Imperial_College/3_October_2008&action=edit 3] [http://2008.igem.org/wiki/index.php?title=Imperial_College/4_October_2008&action=edit 4] [http://2008.igem.org/wiki/index.php?title=Imperial_College/5_October_2008&action=edit 5]
[http://2008.igem.org/wiki/index.php?title=Imperial_College/6_October_2008&action=edit 6] [http://2008.igem.org/Imperial_College/7_October_2008 7] [http://2008.igem.org/wiki/index.php?title=Imperial_College/8_October_2008&action=edit 8] [http://2008.igem.org/Imperial_College/9_October_2008 9] [http://2008.igem.org/wiki/index.php?title=Imperial_College/10_October_2008&action=edit 10] [http://2008.igem.org/wiki/index.php?title=Imperial_College/11_October_2008&action=edit 11] [http://2008.igem.org/wiki/index.php?title=Imperial_College/12_October_2008&action=edit 12]
[http://2008.igem.org/Imperial_College/13_October_2008 13] [http://2008.igem.org/wiki/index.php?title=Imperial_College/14_October_2008&action=edit 14] [http://2008.igem.org/Imperial_College/15_October_2008 15] [http://2008.igem.org/Imperial_College/16_October_2008 16] [http://2008.igem.org/Imperial_College/17_October_2008 17] [http://2008.igem.org/wiki/index.php?title=Imperial_College/18_October_2008&action=edit 18] [http://2008.igem.org/wiki/index.php?title=Imperial_College/19_October_2008&action=edit 19]
[http://2008.igem.org/wiki/index.php?title=Imperial_College/20_October_2008&action=edit 20] [http://2008.igem.org/wiki/index.php?title=Imperial_College/21_October_2008&action=edit 21] [http://2008.igem.org/wiki/index.php?title=Imperial_College/22_October_2008&action=edit 22] [http://2008.igem.org/wiki/index.php?title=Imperial_College/23_October_2008&action=edit 23] [http://2008.igem.org/wiki/index.php?title=Imperial_College/24_October_2008&action=edit 24] [http://2008.igem.org/wiki/index.php?title=Imperial_College/25_October_2008&action=edit 25] [http://2008.igem.org/wiki/index.php?title=Imperial_College/26_October_2008&action=edit 26]
[http://2008.igem.org/wiki/index.php?title=Imperial_College/27_October_2008&action=edit 27] [http://2008.igem.org/wiki/index.php?title=Imperial_College/28_October_2008&action=edit 28] [http://2008.igem.org/wiki/index.php?title=Imperial_College/29_October_2008&action=edit 29] [http://2008.igem.org/wiki/index.php?title=Imperial_College/30_October_2008&action=edit 30] [http://2008.igem.org/wiki/index.php?title=Imperial_College/31_October_2008&action=edit 31]

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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