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Notebook : July 1

Lab work

A - Aerobic/Anaerobic regulation system

Objective : obtaining BBa_K1155000

**1 - Digestion of pSB1C3 plasmid and PCR products : Pndh* by EcoRI/ PstI**

Zhou

We did a PCR amplification. The products were good. After we made digestion by EcoRI/PstI.

Used quantities :

pSB1C3 :
- Plasmid : 4µL
- EcoRI FD : 0.5µL
- PstI FD : 0.5µL
- Buffer FD : 0.8µL
- H2O : 2.2µL

Pndh* :
- Pndh* : 20µL
- EcoRI FD : 0.75µL
- PstI FD : 0.75µL
- Buffer FD : 3µL
- H2O : 5.5µL

We incubate the digestion at 37°C during 3 hours.

2 - Ligation of pSB1C3 and Pndh*

Sheng, Zhou

Used quantities :

Mix A : we mix our digestion mixes :
- Digestion mix of pSB1C3 : 4µL
- Digestion mix of Pndh* : 30µL
- Buffer ligation : 2µL
- H2O : 14µL

Incubate the ligation at 37oC for 1 hour.

Then, we inactivate EcoRI/SpeI activity by ethanol precipitation.

Protocol : Ethanol precipitation

We used 50µL of DNA. At the end, we dilute our DNA in 20µL of H2O.

Finally we did the ligation mix :
- Buffer ligation : 2µL
- Ligase : 1µL
- DNA : 2µL
- H2O : 15µL

We incubate the ligation at 37oC for 1h30.

B - PCB sensor system

Objective : obtaining BphR2 protein

1 - Design of oligos for amplification of BphR2 gene of Pseudomonas pseudoalcaligenes, Pseudomonas oleovorans

Abdou, Sheng, Zhou

We used software gene manager to find the correct oligopeptides for amplification of BphR2 genes in Pseudomonas pseudoalcaligenes, Pseudomonas oleovorans.

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@@ Line 1: / Line 1: @@
 {{Team:Paris_Saclay/incl_debut_generique}}
 ='''Notebook : July 1'''=
 =='''Lab work'''==
-constructing
+==='''A - Aerobic/Anaerobic regulation system'''===
+===='''Objective : obtaining BBa_K1155000'''====
+===='''1 - Digestion of pSB1C3 plasmid and PCR products : Pndh* by EcoRI/ PstI'''====
+Zhou
+We did a PCR amplification. The products were good. After we made digestion by EcoRI/PstI.
+Used quantities :
+* pSB1C3 :
+** Plasmid : 4µL
+** EcoRI FD : 0.5µL
+** PstI FD : 0.5µL
+** Buffer FD : 0.8µL
+** H2O : 2.2µL
+* Pndh* :
+** Pndh* : 20µL
+** EcoRI FD : 0.75µL
+** PstI FD : 0.75µL
+** Buffer FD : 3µL
+** H2O : 5.5µL
+We incubate the digestion at 37°C during 3 hours.
+===='''2 - Ligation of pSB1C3 and Pndh*'''====
+Sheng, Zhou
+Used quantities :
+* Mix A : we mix our digestion mixes :
+** Digestion mix of pSB1C3 : 4µL
+** Digestion mix of Pndh* : 30µL
+** Buffer ligation : 2µL
+** H2O : 14µL
+Incubate the ligation at 37oC for 1 hour.
+* Then, we inactivate EcoRI/SpeI activity by ethanol precipitation.
+Protocol : [[Team:Paris_Saclay/ethanol|Ethanol precipitation]]
+We used 50µL of DNA.
+At the end, we dilute our DNA in 20µL of H2O.
+* Finally we did the ligation mix :
+** Buffer ligation : 2µL
+** Ligase : 1µL
+** DNA : 2µL
+** H2O : 15µL
+We incubate the ligation at 37oC for 1h30.
+==='''B - PCB sensor system'''===
+===='''Objective : obtaining BphR2 protein'''====
+===='''1 - Design of oligos for amplification of BphR2 gene of ''Pseudomonas pseudoalcaligenes'', ''Pseudomonas oleovorans'''''====
+Abdou, Sheng, Zhou
+We used software gene manager to find the correct oligopeptides for amplification of BphR2 genes in ''Pseudomonas pseudoalcaligenes'', ''Pseudomonas oleovorans''.
+{| border="1" align="center"
+|[[Team:Paris Saclay/Notebook/June/28|<big>Previous day</big>]]
+|[[Team:Paris_Saclay/Notebook|<big>Back to calendar</big>]]
+|[[Team:Paris Saclay/Notebook/July/2|<big>Next day</big>]]
+|}
 {{Team:Paris_Saclay/incl_fin}}

Team:Paris Saclay/Notebook/July/1

From 2013.igem.org