IGEM:Paris Bettencourt 2012/Notebooks/suicide group/day by day//2012/07/26: Difference between revisions

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==Transformation result==
==Transformation results==
* Cells of positive control grew but the one we want did not :(
* Cells of positive control grew but the one we want did not :(
=Starting over=
==Liquid culture==
We cultured again the backbones in 10mL LB + 10uL antibiotic
{| style="border-spacing:0;"
| style="border-top:0.002cm solid #000000;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| Name
| style="border-top:0.002cm solid #000000;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| Original name
| style="border-top:0.002cm solid #000000;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| Plate source
| style="border:0.002cm solid #000000;padding:0.097cm;"| Antibiotic
|-
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| PSG.001
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| pSB3C5
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| 04/07/12
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:0.002cm solid #000000;padding:0.097cm;"| Cm
|-
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| PSG.002
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| pSB4K5
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| 18/07/12
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:0.002cm solid #000000;padding:0.097cm;"| Kan
|-
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| PSG.003
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| R0011
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| 09/07/12
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:0.002cm solid #000000;padding:0.097cm;"| Amp
|-
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| PSG.004
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| R0040
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| 04/07/12
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:0.002cm solid #000000;padding:0.097cm;"| Amp
|}
==PCR of Col E2 toxic and immunity gene==
===Recipe (in order)===
{| style="border-spacing:0;"
| style="border-top:0.002cm solid #000000;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| H2O
| style="border:0.002cm solid #000000;padding:0.097cm;"| 37.25 uL
|-
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| 5x Phusion HF Buffer
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:0.002cm solid #000000;padding:0.097cm;"| 10 uL
|-
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| 10 mM dNTPs
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:0.002cm solid #000000;padding:0.097cm;"| 1 uL
|-
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| Primer mix
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:0.002cm solid #000000;padding:0.097cm;"| 1 uL
|-
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| Template DNA
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:0.002cm solid #000000;padding:0.097cm;"| 0.25 uL
|-
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:none;padding:0.097cm;"| Phusion DNA polumerase
| style="border-top:none;border-bottom:0.002cm solid #000000;border-left:0.002cm solid #000000;border-right:0.002cm solid #000000;padding:0.097cm;"| 0.5 uL
|}
*Template DNA used: pSG.012 from 19/07/12
*Primer mix used:
**for pSG.013 (toxic): iSG.003 and iSG.004
**for pSG.014 (immunity): iSG.005 and iSG.006
*Primer mix = 10 uL forward primer + 10 uL reverse primer + 80 uL H2O
===PCR Program===




Revision as of 05:23, 26 July 2012

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Transformation results

  • Cells of positive control grew but the one we want did not :(

Starting over

Liquid culture

We cultured again the backbones in 10mL LB + 10uL antibiotic

Name Original name Plate source Antibiotic
PSG.001 pSB3C5 04/07/12 Cm
PSG.002 pSB4K5 18/07/12 Kan
PSG.003 R0011 09/07/12 Amp
PSG.004 R0040 04/07/12 Amp

PCR of Col E2 toxic and immunity gene

Recipe (in order)

H2O 37.25 uL
5x Phusion HF Buffer 10 uL
10 mM dNTPs 1 uL
Primer mix 1 uL
Template DNA 0.25 uL
Phusion DNA polumerase 0.5 uL
  • Template DNA used: pSG.012 from 19/07/12
  • Primer mix used:
    • for pSG.013 (toxic): iSG.003 and iSG.004
    • for pSG.014 (immunity): iSG.005 and iSG.006
  • Primer mix = 10 uL forward primer + 10 uL reverse primer + 80 uL H2O

PCR Program

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