Team:Paris Saclay/Notebook/August/12

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{{Team:Paris_Saclay/incl_debut_generique}}
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='''Notebook : August 12'''=
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=='''Lab work'''==
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==='''A - Aerobic/Anaerobic regulation system'''===
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===='''Objective : characterize BBa_K1155000'''====
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===='''1 - Digestion of BBa_K1155000 by SpeI/PstI, BBa_K1155007 and BBa_K1155003 by XBaI/PstI'''====
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Anaïs, Nadia, XiaoJing
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Used quantities :
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* BBa_K1155000 :
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** Buffer FD : 5µL
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** H2O : 38µL
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** DNA : 5µL
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** SpeI FD : 1µL
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** PstI FD : 1µL
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* BBa_K1155007 :
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** Buffer FD : 5µL
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** H2O : 23µL
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** DNA : 20µL
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** XBal FD : 1µL
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** PstI FD : 1µL
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* BBa_K1155003 :
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** Buffer FD : 5µL
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** H2O : 33µL
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** DNA : 10µL
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** XBal FD : 1µL
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** PstI FD : 1µL
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We incubated the digestion at 37°C during 15 minutes.
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===='''2 - Electrophoresis to check the digestion of BBa_K1155000 by SpeI/PstI, BBa_K1155007 and BBa_K1155003 by XBalI/PstI'''====
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Nadia
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{|
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| style="width:350px;border:1px solid black;" |[[File:Psgel11208.jpg]]
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| style="width:350px;border:1px solid black;vertical-align:top;" |
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* Well 1 : 6µL DNA Ladder
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* Well 2 : 5µL BBa_K1155000 digested by SpeI/PstI + 1µl of 6X loading dye
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* Well 3 : 5µL BBa_K1155007 digested by XBaI/PstI + 1µl of 6X loading dye
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* Well 4 : 5µL BBa_K1155003 digested by xBaI/PstI + 1µl of 6X loading dye
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* Gel : 0.8%
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|}
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Expected sizes :
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* Pndh* : 111bp
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* RBS_LacZ_Term : 3500 kb
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* RBS_AmilCP_Term : 824 bp
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* pSB1C3 : 2070bp
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{|
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| style="border:1px solid black;padding:5px;background-color:#DEDEDE;" |
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We can't see any band for BBa_K1155000 digestion. The digestion failed. We will do it again. We obtain RBS-LacZ-Term and RBS-AmilCP-Term fragments. The digestion was good. We will purify it.
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|}
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===='''3 - Digestion of BBa_K1155000 by Spe I/Pst I'''====
 +
 +
Anaïs, Nadia
 +
 +
Used quantities :
 +
 +
* Buffer FD : 5µL
 +
* H2O : 38µL
 +
* DNA : 5µL
 +
* SpeI FD : 1µL
 +
* PstI FD : 1µL
 +
 +
We incubate the digestion at 37°C during 15 minutes.
 +
 +
==='''A - Aerobic/Anaerobic regulation system / B - PCB sensor system'''===
 +
 +
===='''Objective : obtaining FNR and BphR2 proteins'''====
 +
 +
===='''1 - Electrophoresis of PCR products : RBS-BphR2 Part I, BphR2 Part I, BphR2 Part II, RBS-FNR Part I, FNR Part I and FNR Part II '''====
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 +
Damir
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{|
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| style="width:350px;border:1px solid black;" |[[File:Psgel21208.jpg]]
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| style="width:350px;border:1px solid black;vertical-align:top;" |
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* Well 1 : 6µL DNA Ladder
 +
* Well 2 : 5µL RBS-BphR2 Part I +1µl of 6X loading dye
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* Well 3 : 5µL BphR2 Part II +1µl of 6X loading dye
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* Well 4 : 5µL FNR Part I +1µl of 6X loading dye
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* Well 5 : 5µL FNR Part II +1µl of 6X loading dye
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* Well 6 : 5µL RBS-FNR Part I +1µl of 6X loading dye
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* Well 7 : 5µL BphR2 Part I +1µl of 6X loading dye
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* Gel : 0.8%
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|}
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Expected size
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* RBS-BphR2 Part I : 197 kb
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* BphR2 Part II : 790 kb
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* FNR Part I : 597 kb
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* FNR Part II : 200 kb
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* RBS-FNR Part I : 615 kb
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* BphR2 Part I : 178 kb
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{|
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| style="border:1px solid black;padding:5px;background-color:#DEDEDE;" |
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We can't see FNR Part I, FNR Part II and BphR2 Part I fragments at the good size. We will make the PCR again. We obtain RBS-BphR2 Part I, BphR2 Part II, RBS-FNR Part I frangments at the right size thanks to the PCR. We will purify it.
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|}
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{{Team:Paris_Saclay/incl_fin}}

Latest revision as of 19:26, 3 October 2013

Navigation Home Team Project Labwork Notebook Human practices

Contents

Notebook : August 12

Lab work

A - Aerobic/Anaerobic regulation system

Objective : characterize BBa_K1155000

1 - Digestion of BBa_K1155000 by SpeI/PstI, BBa_K1155007 and BBa_K1155003 by XBaI/PstI

Anaïs, Nadia, XiaoJing

Used quantities :

  • BBa_K1155000 :
    • Buffer FD : 5µL
    • H2O : 38µL
    • DNA : 5µL
    • SpeI FD : 1µL
    • PstI FD : 1µL
  • BBa_K1155007 :
    • Buffer FD : 5µL
    • H2O : 23µL
    • DNA : 20µL
    • XBal FD : 1µL
    • PstI FD : 1µL
  • BBa_K1155003 :
    • Buffer FD : 5µL
    • H2O : 33µL
    • DNA : 10µL
    • XBal FD : 1µL
    • PstI FD : 1µL

We incubated the digestion at 37°C during 15 minutes.

2 - Electrophoresis to check the digestion of BBa_K1155000 by SpeI/PstI, BBa_K1155007 and BBa_K1155003 by XBalI/PstI

Nadia

Psgel11208.jpg
  • Well 1 : 6µL DNA Ladder
  • Well 2 : 5µL BBa_K1155000 digested by SpeI/PstI + 1µl of 6X loading dye
  • Well 3 : 5µL BBa_K1155007 digested by XBaI/PstI + 1µl of 6X loading dye
  • Well 4 : 5µL BBa_K1155003 digested by xBaI/PstI + 1µl of 6X loading dye
  • Gel : 0.8%

Expected sizes :

  • Pndh* : 111bp
  • RBS_LacZ_Term : 3500 kb
  • RBS_AmilCP_Term : 824 bp
  • pSB1C3 : 2070bp

We can't see any band for BBa_K1155000 digestion. The digestion failed. We will do it again. We obtain RBS-LacZ-Term and RBS-AmilCP-Term fragments. The digestion was good. We will purify it.

3 - Digestion of BBa_K1155000 by Spe I/Pst I

Anaïs, Nadia

Used quantities :

  • Buffer FD : 5µL
  • H2O : 38µL
  • DNA : 5µL
  • SpeI FD : 1µL
  • PstI FD : 1µL

We incubate the digestion at 37°C during 15 minutes.

A - Aerobic/Anaerobic regulation system / B - PCB sensor system

Objective : obtaining FNR and BphR2 proteins

1 - Electrophoresis of PCR products : RBS-BphR2 Part I, BphR2 Part I, BphR2 Part II, RBS-FNR Part I, FNR Part I and FNR Part II

Damir

Psgel21208.jpg
  • Well 1 : 6µL DNA Ladder
  • Well 2 : 5µL RBS-BphR2 Part I +1µl of 6X loading dye
  • Well 3 : 5µL BphR2 Part II +1µl of 6X loading dye
  • Well 4 : 5µL FNR Part I +1µl of 6X loading dye
  • Well 5 : 5µL FNR Part II +1µl of 6X loading dye
  • Well 6 : 5µL RBS-FNR Part I +1µl of 6X loading dye
  • Well 7 : 5µL BphR2 Part I +1µl of 6X loading dye
  • Gel : 0.8%

Expected size

  • RBS-BphR2 Part I : 197 kb
  • BphR2 Part II : 790 kb
  • FNR Part I : 597 kb
  • FNR Part II : 200 kb
  • RBS-FNR Part I : 615 kb
  • BphR2 Part I : 178 kb

We can't see FNR Part I, FNR Part II and BphR2 Part I fragments at the good size. We will make the PCR again. We obtain RBS-BphR2 Part I, BphR2 Part II, RBS-FNR Part I frangments at the right size thanks to the PCR. We will purify it.


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