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Notebook : July 4

Lab work

A - Aerobic/Anaerobic regulation system

Objective : obtaining BBa_K1155000

**1 - Estimation of Pndh* Colony PCR size fragments**

Abdou, Anaïs, Sheng

We used software gene manager to find the correct size of our fragments.

Estimated size :

VF/VR primer -> Plasmid without Pndh* size : 277bp
VF/Pfnr_down -> Plasmid with Pndh* size : 276bp
Pfnr_Up/VR -> Plasmid with Pndh* size : 311bp

2 - Electrophoresis of Colony PCR products

Zhou

Well 1 to 4 : 5µL of PCR products with VF/VR primers+1µL of 6X loading dye
Well 5 to 6 : 5µL of PCR product withVF/Pfnr_Down primers+1µL of 6X loading dye
Well 7 : 6µL of DNA Ladder
Well 8 to 9 : 5µL of PCR product withVF/Pfnr_Down primers+1µL of 6X loading dye
Well 10 to 13 : 5µL of PCR products with Pfnr_Up/VR primers+1µL of 6X loading dye
Well 14 : 6µL of DNA Ladder
Gel : 1.5%

We obtain fragment at the right size in wells : 3, 4 , 8 , 9 12 and 13. We will digest our biobrick to confirm that Pndh* insert pSB1C3 correctly.

3 - Stock of BBa_K1155000

Zhou

Used quantities :

BBa_K1155000 confirmed : 1 mL
Glycerol : 500µL glycerol.

We stocked them at -20°C.

4 - Extraction of BBa_K1155000 from DH5α

Anaïs, Sheng

Protocol : Hight copy plamid extraction

**5 - Digestion of BBa_K1155000 by NotI, MluI, HpaI to check good insertion of Pndh* in pSB1C3**

Abdou

Used quantities :

BBa_K1155000 : 2µL
Bufer oranger : 2µL
NotI or MluI or HpaI : 0.5µL
H2O : 15.5µL

We let the digestion 1h30 at 37°C.

6 - Culture of BBa_K1155000

Anaïs, Sheng

We made 2 cultures of bacterias transformed with BBa_K1155000 that show a fragments at the right size at electrophoresis.

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@@ Line 7: / Line 7: @@
 ==='''A - Aerobic/Anaerobic regulation system'''===
-===='''Objective : obtaining Bba_K1155000'''====
+===='''Objective : obtaining BBa_K1155000'''====
 ===='''1 - Estimation of Pndh* Colony PCR size fragments'''====
@@ Line 38: / Line 38: @@
 {|
 | style="border:1px solid black;padding:5px;background-color:#DEDEDE;" |
-We obtain fragment at the right size in wells : 3, 4 , 8 , 9 12 and 13. We will digest our biobrick to confirm that Pndh* insert PSB1C3 correctly.
+We obtain fragment at the right size in wells : 3, 4 , 8 , 9 12 and 13. We will digest our biobrick to confirm that Pndh* insert pSB1C3 correctly.
 |}
-===='''3 - Stock of Bba_K1155000'''====
+===='''3 - Stock of BBa_K1155000'''====
 Zhou
 Used quantities :
-* Bba_K1155000 confirmed : 1 mL
+* BBa_K1155000 confirmed : 1 mL
 * Glycerol : 500µL glycerol.
 We stocked them at -20°C.
-===='''4 - Extraction of Bba_K1155000 from DH5α'''====
+===='''4 - Extraction of BBa_K1155000 from DH5α'''====
 Anaïs, Sheng
@@ Line 57: / Line 57: @@
 Protocol : [[Team:Paris_Saclay/extraction|Hight copy plamid extraction]]
-===='''5 - Digestion of Bba_K1155000 by NotI, MluI, HpaI to check good insertion of Pndh* in PSB1C3'''====
+===='''5 - Digestion of BBa_K1155000 by NotI, MluI, HpaI to check good insertion of Pndh* in pSB1C3'''====
 Abdou
 Used quantities :
-* Bba_K1155000 : 2µL
+* BBa_K1155000 : 2µL
 * Bufer oranger : 2µL
 * NotI or MluI or HpaI : 0.5µL
@@ Line 69: / Line 69: @@
 We let the digestion 1h30 at 37°C.
-===='''6 - Culture of Bba_K1155000'''====
+===='''6 - Culture of BBa_K1155000'''====
 Anaïs, Sheng
-We made 2 cultures of bacterias transformed with Bba_K1155000 that show a fragments at the right size at electrophoresis.
+We made 2 cultures of bacterias transformed with BBa_K1155000 that show a fragments at the right size at electrophoresis.

Team:Paris Saclay/Notebook/July/4

From 2013.igem.org