Team:Paris Saclay/Labwork

From 2013.igem.org

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07/03/13
07/03/13
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* PCR on colonies on clones obtained after the transformation to check good insertion of Pndh* in pSB1C3
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* PCR on colonies obtained after the transformation to check good insertion of Pndh* in pSB1C3
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* New culture of Bba_K1155000
+
* Colonies assayed by PCR streaked on plates and used for liquid cultures
07/04/13
07/04/13
* Estimation of the expected PCR size fragments using bio-informatic tools
* Estimation of the expected PCR size fragments using bio-informatic tools
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* Gel Electrophoresis of PCR products obtained the 07/03/13 [[File:Psright.png|20px]]
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* Gel Electrophoresis of PCR products obtained the day before [[File:Psright.png|20px]]
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* Stock of Bba_K1155000
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* Glycerol Stocks of the clones obtained
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* Culture of Bba_K1155000
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* DNA Extraction of the plasmids from DH5α
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* Extraction of Bba_K1155000 from DH5α
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* Digestion of the plasmids by NotI, MluI, HpaI to check with a different method the good insertion of Pndh* in pSB1C3
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* Digestion of Bba_K1155000 by NotI, MluI, HpaI to check good insertion of Pfnr in PSB1C3 again
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* New liquid culture of the correct clone
07/05/13
07/05/13
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* Electrophoresis of the digestion of Bba_K1155000 by NotI, HpaI, MluI [[File:Pswrong.png|20px]]
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* Electrophoresis of the digestion of the correct cloned , now named Bba_K1155000 by NotI, HpaI, MluI [[File:Pswrong.png|20px]]
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* Extraction of Bba_K1155000 from DH5α
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* Extraction of plasmidic DNA of Bba_K1155000 from DH5α for sequencing
07/15/13
07/15/13
* Sequense analysis of Bba_K1155000 [[File:Psright.png|20px]]
* Sequense analysis of Bba_K1155000 [[File:Psright.png|20px]]
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WE OBTAIN OUR BIOBRICK.
+
WE OBTAINED OUR BIOBRICK.
=='''Bba_K1155001'''==
=='''Bba_K1155001'''==

Revision as of 16:58, 27 September 2013

Labwork Overview Biobricks Protocols Safety

Contents

Labwork overview

Bba_K1155000

Engineered promoter region of E. coli ndh gene that allows repression of the expression of the downstream gene in anaerobic conditions

07/01/13

  • Digestion of pSB1C3 plasmid and Pndh* PCR products by EcoRI/ PstI
  • Ligation of pSB1C3 and Pndh*

07/02/13

  • Tranformation of E. coli DH5α with the ligation solution Psright.png

07/03/13

  • PCR on colonies obtained after the transformation to check good insertion of Pndh* in pSB1C3
  • Colonies assayed by PCR streaked on plates and used for liquid cultures

07/04/13

  • Estimation of the expected PCR size fragments using bio-informatic tools
  • Gel Electrophoresis of PCR products obtained the day before Psright.png
  • Glycerol Stocks of the clones obtained
  • DNA Extraction of the plasmids from DH5α
  • Digestion of the plasmids by NotI, MluI, HpaI to check with a different method the good insertion of Pndh* in pSB1C3
  • New liquid culture of the correct clone

07/05/13

  • Electrophoresis of the digestion of the correct cloned , now named Bba_K1155000 by NotI, HpaI, MluI Pswrong.png
  • Extraction of plasmidic DNA of Bba_K1155000 from DH5α for sequencing

07/15/13

  • Sequense analysis of Bba_K1155000 Psright.png

WE OBTAINED OUR BIOBRICK.

Bba_K1155001

07/09/13

  • PCR purification of BphR1
  • Digestion PCR products : BphR1 and digestion of PSB1C3
  • Denaturation of EcoRI/PstI used for the digestion of PCR products : BphR1 and PSB1C3
  • Ligation of BphR1 and PSB1C3

07/10/13

  • Transformation of ligation of PSB1C3 and BphR1 Psright.png

07/11/13

  • Colony PCR of Bba_K1155001 in DH5α to check good insertion of BphR1 in PSB1C3
  • Electrophoresis of Colony PCR products : Bba_K1155001 in PSB1C3 Psright.png

07/12/13

  • Stock of clones 5, 6 for Bba_K1155001
  • Extraction of clones 5, 6 for Bba_K1155001 from DH5α

07/15/13

  • Digestion of Bba_K1155001 by EcoRI/PstI and SacII to check the size of fragments
  • PCR of digestion of Bba_K1155001

07/16/13

  • Electrophoresis of the digestion of Bba_K1155001 Psright.png

07/17/13

  • Stock of Bba_K1155001
  • Extraction of Bba_K1155001 from DH5α

07/24/13

  • Sequence analysis of Bba_K1155001 Psright.png

WE OBTAIN OUR BIOBRICK.


Bba_K1155002

07/09/13

  • PCR purification of BphA1
  • Digestion PCR products : BphA1 and digestion of PSB1C3
  • Denaturation of EcoRI/PstI used for the digestion of PCR products : BphA1 and PSB1C3
  • Ligation of BphA1 and PSB1C3

07/10/13

  • Transformation of ligation of PSB1C3 and BphA1 Psright.png

07/11/13

  • Colony PCR of Bba_K1155002 in DH5α to check good insertion of BphA1 in PSB1C3
  • Electrophoresis of Colony PCR products : Bba_K1155002 in PSB1C3 Psright.png

07/12/13

  • Stock of clones 5, 6, 7, 8 for Bba_K1155002
  • Streak of Bba_K1155002
  • Extraction of clones 5, 6, 7, 8 for Bba_K1155002 from DH5α

07/15/13

  • Digestion of Bba_K1155002 by EcoRI/PstI and SacII to check the size of fragments
  • PCR of digestion of Bba_K1155002

07/16/13

  • Electrophoresis of the digestion of Bba_K1155002 Pswrong.png

07/25/13

  • Ligation of BphA1 in PSB1C3

07/26/13

  • Tranformation of Bba_K1155002 in DH5α Psright.png

07/30/13

  • Colony PCR of Bba_K1155002 in DH5α
  • Electrophoresis of the Colony PCR of Bba_K1155002 in DH5α Psright.png


Bba_K1155003

07/02/13

  • Tranformation of Bba_K592009 in DH5α Psright.png

07/03/13

  • New culture of Bba_K592009

07/05/13

  • Extraction of Bba_K592009 from DH5α

07/08/13

  • Digestion of Bba_K592009 by NotI, XhoI, EcoRI, EcoRI/PstI
  • Electrophoresis of the digestion of Bba_K592009 by NotI, XhoI, EcoRI, EcoRI/PstI Psright.png

07/12/13

  • Transformation of Bba_B0015, Bba_B0017 Psright.png
  • Design of RBS-AmilCP oligos

07/15/13

  • Liquid culture of Bba_B0015, Bba_B0017 Psright.png

07/16/13

  • Extraction of Bba_B0015, Bba_B0017 from DH5α
  • Digestion of Bba_B0015, Bba_B0017, Bba_I732019 by EcoRI/PstI

07/18/13

  • PCR of Bba_B0015, Bba_B0017 digested by EcoRI/PstI

07/19/13

  • Electrophoresis of Colony PCR products : Bba_B0015, BBa_B0017 Psright.png
  • Gel purification of the electrophoresis of Colony PCR products : Bba_B0015, BBa_B0017 Psright.png

07/22/13

  • Digestion of Bba_K592009 by SpeI/PstI, Bba_B0015 and Bba_B0017 by PstI/XbaI
  • Denaturation of SpeI/PstI and PstI/XbaI used for the digestion of Bba_K592009, Bba_B0015 and Bba_B0017

07/23/13

  • Electrophoresis of the digestion of Bba_K592009 by SpeI/PstI Pswrong.png
  • Electrophoresis of the digestion of Bba_B0015, Bba_B0017 by XbaI/PstI Psright.png
  • Digestion of Bba_K592009 by SpeI/PstI
  • Electrophoresis to check the digestion of Bba_K592009 by SpeI/PstI Pswrong.png
  • Liquid culture of Bba_K592009 in DH5α Psright.png

07/24/13

  • Extraction of Bba_K592009 from DH5α

07/25/13

  • Gel purification of Term
  • Electrophoresis to check the gel purification of Term Psright.png
  • PCR of Bba_K592209 with RBS-Amil CP primer and suffix : PSB1C3
  • Digestion of Colony PCR products : RBS-Amil CP by EcoRI/SpeI

07/26/13

  • Denaturation of EcoRI/SpeI used for the digestion of PCR products : RBS-AmilCP
  • Ligation of RBS-AmilCP and Term-PSB1C3

07/30/13

  • Tranformation of Bba_K1155003 in DH5α


Bba_K1155004

07/25/13

  • Colony PCR of NirB
  • Electrophoresis to check the colony PCR of NirB Pswrong.png
  • Colony PCR of NirB

07/26/13

  • Electrophoresis of PCR products : NirB Pswrong.png
  • PCR of NirB
  • Electrophoresis of PCR products : NirB Psright.png
  • Gel purification of PCR products : NarK Psright.png
  • Digestion of PCR products : NarK by EcoRI/PstI

07/30/13

  • Denaturation of EcoRI/SpeI used of the digestion of PCR products : NirB

07/31/13

  • Digestion of PSB1C3 by EcoRI/PstI'
  • Denaturation of EcoRI/PstI used for the digestion of PSB1C3
  • Ligation of NirB in PSB1C3
  • Electrophoresis to check the ligation of NirB in PSB1C3
  • Transformation of Bba_K1155004


Bba_K1155005

07/25/13

  • Colony PCR of nag
  • Electrophoresis to check the colony PCR of NarG Pswrong.png
  • Colony PCR of NarG

07/26/13

  • Electrophoresis of PCR products : NarG Pswrong.png
  • PCR of NarG
  • Electrophoresis of PCR products : NarG Psright.png
  • Gel purification of PCR products : NarG Psright.png
  • Digestion of PCR products : NarG by EcoRI/PstI

07/30/13

  • Denaturation of EcoRI/SpeI used of the digestion of PCR products : NarG

07/31/13

  • Digestion of PSB1C3 by EcoRI/PstI'
  • Denaturation of EcoRI/PstI used for the digestion of PSB1C3
  • Ligation of NarG in PSB1C3
  • Electrophoresis to check the ligation of NarG in PSB1C3
  • Transformation of Bba_K1155005


Bba_K1155006

07/25/13

  • Colony PCR of NarK
  • Electrophoresis to check the colony PCR of NarK Pswrong.png
  • Colony PCR of NarK

07/26/13

  • Electrophoresis of PCR products : NarK Pswrong.png
  • PCR of NarK
  • Electrophoresis of PCR products : NarK Psright.png
  • Gel purification of PCR products : NirB Psright.png
  • Digestion of PCR products : NirB by EcoRI/PstI

07/30/13

  • Denaturation of EcoRI/SpeI used of the digestion of PCR products : NarK

07/31/13

  • Digestion of PSB1C3 by EcoRI/PstI'
  • Denaturation of EcoRI/PstI used for the digestion of PSB1C3
  • Ligation of NarK in PSB1C3
  • Electrophoresis to check the ligation of NarK in PSB1C3
  • Transformation of Bba_K1155006


Bba_K1155007

07/02/13

  • Tranformation of Bba_I732017 in DH5α Psright.png

07/03/13

  • New culture of Bba_I732017

07/05/13

  • Extraction of Bba_I732017 from DH5α

07/08/13

  • Digestion of Bba_I732017 by NotI, XhoI, EcoRI, EcoRI/PstI
  • Electrophoresis of the digestion of Bba_I732017 by NotI, XhoI, EcoRI, EcoRI/PstI Psright.png

07/10/13

  • Transformation of Bba_I732019 in DH5α Pswrong.png

07/12/13

  • Transformation of Bba_I732019, Bba_B0015, Bba_B0017 Psright.png

07/15/13

  • Liquid culture of Bba_B0015, Bba_B0017, Bba_I732019 Psright.png

07/16/13

  • Extraction of Bba_B0015, Bba_B0017, Bba_I732019 from DH5α
  • Digestion of Bba_B0015, Bba_B0017, Bba_I732019 by EcoRI/PstI

07/17/13

  • Electrophoresis of digestions of Bba_B0015, Bba_B0017 by EcoRI/PstI Psright.png
  • Electrophoresis of digestions of Bba_I732019 by EcoRI/PstI Pswrong.png

07/18/13

  • PCR of Bba_B0015, Bba_B0017 digested by EcoRI/PstI

07/19/13

  • Electrophoresis of Colony PCR products : Bba_B0015, BBa_B0017 Psright.png
  • Gel purification of the electrophoresis of Colony PCR products : Bba_B0015, BBa_B0017 Psright.png

07/22/13

  • Digestion of Bba_I732017 by EcoRI/SpeI, Bba_B0015 and Bba_B0017 by PstI/XbaI
  • Denaturation of EcoRI/SpeI and PstI/XbaI used for the digestion of Bba_I732017, Bba_K502009, Bba_B0015 and Bba_B0017

07/23/13

  • Electrophoresis of the digestion of Bba_I732017 by EcoRI/SpeI, Bba_B0015, Bba_B0017 by XbaI/PstI Psright.png
  • Gel purification of digestion products : RBS-LacZ
  • Electrophoresis to check the gel purification of digestion products : RBS-LacZ Pswrong.png
  • Digestion of Bba_I732017 by EcoRI/Pst
  • Electrophoresis to check the gel digestion of Bba_I732017 by EcoRI/SpeI Pswrong.png
  • Liquid culture of Bba_I732017 in DH5α Psright.png

07/24/13

  • Extraction of Bba_I732017 from DH5α

07/25/13

  • Gel purification of Term
  • Electrophoresis to check the gel purification of Term Psright.png

07/30/13

  • Digestion of Bba_I732017 by EcoRI/PstI
  • Electrophoresis to check the digestion of Bba_I732017 by EcoRI/PstI Psright.png
  • Gel purification of digestion of Bba_I732017 by EcoRI/PstI Psright.png
  • Digestion of Bba_I732017 by EcoRI/PstI
  • Electrophoresis to check the digestion of Bba_I732017 by EcoRI/SpeI Psright.png
  • Gel purification of digestion of Bba_I732017 by EcoRI/PstI Pswrong.png

07/31/13

  • Digestion of Bba_I732017 by EcoRI/SpeI
  • Electrophoresis to check the digestion of Bba_I732017 by EcoRI/SpeI Psright.png
  • Gel purification of digestion of RBS-LacZ Pswrong.png


Bba_K1155008

07/30/13

  • PCR of : FNR Part I, FNR Part II, RBS-FNR Part I
  • Electrophoresis of the PCR products : FNR Part I, FNR Part II, RBS-FNR Part I Psright.png


Bba_K1155009

07/01/13

  • Design of oligos for amplification of BphR2 gene of Pseudomonas pseudoalcaligenes, Pseudomonas oleovorans

07/09/13

  • PCR purification of BphR2
  • Digestion PCR products : BphR2 and digestion of PSB1C3
  • Denaturation of EcoRI/PstI used for the digestion of PCR products : BphR2 and PSB1C3
  • Ligation of BphR2 and PSB1C3

07/10/13

  • Transformation of ligation of PSB1C3 and BphR2 Psright.png

07/11/13

  • Colony PCR of BphR2 in PSB1C3 in DH5α to check good insertion of BphR2 in PSB1C3
  • Electrophoresis of Colony PCR products : BphR2 in PSB1C3 Psright.png

07/12/13

  • Stock of clones 3, 4 for BphR2
  • Streak of BphR2
  • Extraction of clones 3, 4 for BphR2 from DH5α

07/15/13

  • Digestion of BphR2 in PSB1C3 by EcoRI/PstI and SacII to check the size of fragments
  • PCR of digestion of BphR2 in PSB1C3

07/16/13

  • Electrophoresis of the digestion of BphR2 in PSB1C3 Pswrong.png

07/30/13

  • PCR of : BphR2 Part I, BphR2 Part II, RBS-BphR2 Part I
  • Electrophoresis of the PCR products : BphR2 Part I, BphR2 Part II Psright.png
  • Electrophoresis of the PCR products : RBS-BphR2 Part I Pswrong.png
  • PCR of : RBS-BphR2 Part I
  • Gel purification of PCR of : RBS-BphR2 Part I Pswrong.png


PSB3K3

07/09/13

  • Transformation of Bba_J04450 in DH5α Psright.png

07/10/13

  • Mini and maxi preparation of the culture of Bba_J04450 Psright.png

07/11/13

  • Extraction of Bba_J04450 from DH5α
  • Digestion of Bba_J04450 to chek the size for the plasmid
  • Electrophoresis of the digestion of Bba_J04450 Psright.png

07/16/13

  • Digestion of PSB3K3 by EcoRI/PstI
  • Electrophoresis of the digestion of PSB3K3 by EcoRI/PstI Psright.png
  • Gel purification of electrophoresis of the digestion of PSB3K3 by EcoRI/PstI Psright.png

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