Team:Paris Saclay/Notebook/July/3

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{{Team:Paris_Saclay/incl_debut_generique}}
{{Team:Paris_Saclay/incl_debut_generique}}
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='''Notebook : July 3'''=
='''Notebook : July 3'''=
=='''Lab work'''==
=='''Lab work'''==
-
constructing
 
 +
==='''A - Aerobic/Anaerobic regulation system'''===
 +
===='''Objective : obtaining BBa_K1155000'''====
 +
 +
===='''1 - Colony PCR of BBa_K1155000 to check good insertion of Pndh* in pSB1C3'''====
 +
 +
Abdou, Sheng, Zhou
 +
 +
{|
 +
| style="border:1px solid black;padding:5px;background-color:#DE;" |
 +
Tranformation of 07/02/13 works. We will do a PCR colony.
 +
|}
 +
 +
Colonies count for BBa_K1155000 :
 +
 +
* Standard concentration :
 +
** BBa_K1155000 : 0 colony
 +
 +
* High concentration :
 +
** BBa_K1155000 : 2 colonies
 +
[[File:Ps0307jour.jpg|300px]]
<br>
<br>
 +
<center>[[File:PSprimer07.jpg|right|250px]]</center>
 +
<br>
 +
'''[https://2013.igem.org/Team:Paris_Saclay/Primers Primers] and PCR :'''
 +
<p>'''VF2, VR, Pfnr_up, Pfnr_down are four oligos that we used for plasmid amplification. We used tree combinaisons VF/VR, VF/Pfnr_Down, Pfnr_Up/VR.'''
 +
'''If the promoter Pndh* insert pSB1C3 plasmid successfully, tree fragments with specific size will be amplified.''' </p>
 +
'''
 +
Protocol'''
 +
 +
We resuspend each colony with 20µL of H2O.
 +
 +
Used quantities :
 +
* DNA : 2µL
 +
* Mix : (it was divided in tubes for 4 different colonies for each oligo combinaison with 23µL of mix in each tube)
 +
** VF or Pfnr_Up : 6µL
 +
** VR or Pfnr_Down : 6µL
 +
** dNTP 10mM : 6µL
 +
** Buffer Dream Taq : 30µL
 +
** Dream Taq : 6µL
 +
** H2O : 246µL
 +
 +
[[File:PSPCR0307.jpg|400px]]
 +
 +
===='''2 - Culture of BBa_K1155000 '''====
 +
 +
Sheng
 +
 +
We made new cultures by streaking 2 colonies of 07/02/13 culture of BBa_K1155000. We also did liquid culture of each one. 
 +
We incubate culture at 37°C.
 +
 +
===='''Objective : obtaining BBa_K1155003, BBa_K1155007'''====
 +
 +
===='''1 - Culture of BBa_I732017, BBa_K592009 '''====
 +
 +
Sheng
 +
 +
{|
 +
| style="border:1px solid black;padding:5px;background-color:#DE;" |
 +
Tranformation of 07/02/13 works. We will do new cultures.
 +
|}
 +
 +
We made new cultures by streaking 2 colonies of 07/02/13 culture of BBa_I732017, BBa_K592009. We also did liquid culture of each one. 
 +
We incubate culture at 37°C.
 +
 +
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Latest revision as of 00:51, 5 October 2013

Navigation Home Team Project Labwork Notebook Human practices

Contents

Notebook : July 3

Lab work

A - Aerobic/Anaerobic regulation system

Objective : obtaining BBa_K1155000

1 - Colony PCR of BBa_K1155000 to check good insertion of Pndh* in pSB1C3

Abdou, Sheng, Zhou

Tranformation of 07/02/13 works. We will do a PCR colony.

Colonies count for BBa_K1155000 :

  • Standard concentration :
    • BBa_K1155000 : 0 colony
  • High concentration :
    • BBa_K1155000 : 2 colonies

Ps0307jour.jpg


PSprimer07.jpg


Primers and PCR :

VF2, VR, Pfnr_up, Pfnr_down are four oligos that we used for plasmid amplification. We used tree combinaisons VF/VR, VF/Pfnr_Down, Pfnr_Up/VR. If the promoter Pndh* insert pSB1C3 plasmid successfully, tree fragments with specific size will be amplified.

Protocol

We resuspend each colony with 20µL of H2O.

Used quantities :

  • DNA : 2µL
  • Mix : (it was divided in tubes for 4 different colonies for each oligo combinaison with 23µL of mix in each tube)
    • VF or Pfnr_Up : 6µL
    • VR or Pfnr_Down : 6µL
    • dNTP 10mM : 6µL
    • Buffer Dream Taq : 30µL
    • Dream Taq : 6µL
    • H2O : 246µL

PSPCR0307.jpg

2 - Culture of BBa_K1155000

Sheng

We made new cultures by streaking 2 colonies of 07/02/13 culture of BBa_K1155000. We also did liquid culture of each one. We incubate culture at 37°C.

Objective : obtaining BBa_K1155003, BBa_K1155007

1 - Culture of BBa_I732017, BBa_K592009

Sheng

Tranformation of 07/02/13 works. We will do new cultures.

We made new cultures by streaking 2 colonies of 07/02/13 culture of BBa_I732017, BBa_K592009. We also did liquid culture of each one. We incubate culture at 37°C.


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