User:DavidC/13 September 2009

From 2009.igem.org

(Difference between revisions)
Line 16: Line 16:
=== Sunday the 13th ===
=== Sunday the 13th ===
-
==== Restriction ====
+
==== Restriction digest ====
-
==== Ligation of C0012 with B0014: ====
+
==== Ligation between BBa_C0012 and BBa_B0014: ====
-
B0014 (=2,41µg/µL) restriction digest by EcoRI and XbaI (3284bp): <br>
+
 
 +
BBa_B0014 (=2,41µg/µL) restriction digest by EcoRI and XbaI (3284bp): <br>
DNA (10µg final)= 4,2µL <br>
DNA (10µg final)= 4,2µL <br>
Line 26: Line 27:
Eco R1 = 1µL <br>
Eco R1 = 1µL <br>
Incubation 1h at 37°C. <br>
Incubation 1h at 37°C. <br>
 +
DNA purification with a nucleospin (macherey-nagel) . <br>
DNA purification with a nucleospin (macherey-nagel) . <br>
 +
DNA = 50µL <br>
DNA = 50µL <br>
Buffer 2 (NEB) = 6µL <br>
Buffer 2 (NEB) = 6µL <br>
Line 32: Line 35:
H20 = 2,4µL <br>
H20 = 2,4µL <br>
Xba 1 = 1µL <br>
Xba 1 = 1µL <br>
-
 
Incubation 1h at 37°C. <br><br>
Incubation 1h at 37°C. <br><br>
C0012 (1,55µg/µL) restriction digest by EcoRI and SpeI (1128bp): <br>
C0012 (1,55µg/µL) restriction digest by EcoRI and SpeI (1128bp): <br>
 +
DNA (10µg final) = 6,4µL <br>
DNA (10µg final) = 6,4µL <br>
Buffer Eco R1 (NEB) = 5µL <br>
Buffer Eco R1 (NEB) = 5µL <br>
Line 41: Line 44:
BSA = 0,5µL <br>
BSA = 0,5µL <br>
Eco R1 (NEB) = 1µL <br>
Eco R1 (NEB) = 1µL <br>
-
 
Spe 1 (NEB) = 1µL <br>
Spe 1 (NEB) = 1µL <br>
-
 
Incubation 1h at 37°C. <br>
Incubation 1h at 37°C. <br>
==== TAKARA Enzyme testing ====
==== TAKARA Enzyme testing ====
-
==== Ligation of C0012 with B0014: ====
+
==== Ligation between BBa_C0012 and BBa_B0014: ====
-
B0014 (= 2,405µg/µL) restriction digest with EcoRI and XbaI (3284bp): <br>
+
 
 +
B0014 (= 2,405µg/µL) restriction digest with EcoRI and XbaI (3128bp): <br>
 +
 
ADN (10µg final) = 4,2µL <br>
ADN (10µg final) = 4,2µL <br>
Buffer M (TAKARA) = 5µL <br>
Buffer M (TAKARA) = 5µL <br>
Line 55: Line 58:
Eco R1 = 1µL <br>
Eco R1 = 1µL <br>
Xba 1 = 1µL <br>
Xba 1 = 1µL <br>
-
 
Incubation 1h à 37°C. <br>
Incubation 1h à 37°C. <br>
Line 64: Line 66:
Ladder fermentas 1 Kb. <br>
Ladder fermentas 1 Kb. <br>
-
Samples: BBa_B0014, BBa_C0012, BBa_B0014.<br>
+
Samples:  
-
 
+
-
[[Image:S1309.png]]
+
==== DNA purification ====
==== DNA purification ====
Kit Qiagen “gel extraction kit”, final volume = 50µL. <br>
Kit Qiagen “gel extraction kit”, final volume = 50µL. <br>

Revision as of 03:12, 21 October 2009

framless



August
MTWTFSS
          [http://2009.igem.org/User:DavidC/1_August_2009 1] [http://2009.igem.org/User:DavidC/2_August_2009 2]
[http://2009.igem.org/User:DavidC/3_August_2009 3] [http://2009.igem.org/User:DavidC/4_August_2009 4] [http://2009.igem.org/User:DavidC/5_August_2009 5] [http://2009.igem.org/User:DavidC/6_August_2009 6] [http://2009.igem.org/User:DavidC/7_August_2009 7] [http://2009.igem.org/User:DavidC/8_August_2009 8] [http://2009.igem.org/User:DavidC/9_August_2009 9]
[http://2009.igem.org/User:DavidC/10_August_2009 10] [http://2009.igem.org/User:DavidC/11_August_2009 11] [http://2009.igem.org/User:DavidC/12_August_2009 12] [http://2009.igem.org/User:DavidC/13_August_2009 13] [http://2009.igem.org/User:DavidC/14_August_2009 14] [http://2009.igem.org/User:DavidC/15_August_2009 15] [http://2009.igem.org/User:DavidC/16_August_2009 16]
[http://2009.igem.org/User:DavidC/17_August_2009 17] [http://2009.igem.org/User:DavidC/18_August_2009 18] [http://2009.igem.org/User:DavidC/19_August_2009 19] [http://2009.igem.org/User:DavidC/20_August_2009 20] [http://2009.igem.org/User:DavidC/21_August_2009 21] [http://2009.igem.org/User:DavidC/22_August_2009 22] [http://2009.igem.org/User:DavidC/23_August_2009 23]
[http://2009.igem.org/User:DavidC/24_August_2009 24] [http://2009.igem.org/User:DavidC/25_August_2009 25] [http://2009.igem.org/User:DavidC/26_August_2009 26] [http://2009.igem.org/User:DavidC/27_August_2009 27] [http://2009.igem.org/User:DavidC/28_August_2009 28] [http://2009.igem.org/User:DavidC/29_August_2009 29] [http://2009.igem.org/User:DavidC/30_August_2009 30]
[http://2009.igem.org/User:DavidC/31_August_2009 31]
September
MTWTFSS
  [http://2009.igem.org/User:DavidC/1_September_2009 1] [http://2009.igem.org/User:DavidC/2_September_2009 2] [http://2009.igem.org/User:DavidC/3_September_2009 3] [http://2009.igem.org/User:DavidC/4_September_2009 4] [http://2009.igem.org/User:DavidC/5_September_2009 5] [http://2009.igem.org/User:DavidC/6_September_2009 6]
[http://2009.igem.org/User:DavidC/7_September_2009 7] [http://2009.igem.org/User:DavidC/8_September_2009 8] [http://2009.igem.org/User:DavidC/9_September_2009 9] [http://2009.igem.org/User:DavidC/10_September_2009 10] [http://2009.igem.org/User:DavidC/11_September_2009 11] [http://2009.igem.org/User:DavidC/12_September_2009 12] [http://2009.igem.org/User:DavidC/13_September_2009 13]
[http://2009.igem.org/User:DavidC/14_September_2009 14] [http://2009.igem.org/User:DavidC/15_September_2009 15] [http://2009.igem.org/User:DavidC/16_September_2009 16] [http://2009.igem.org/User:DavidC/17_September_2009 17] [http://2009.igem.org/User:DavidC/18_September_2009 18] [http://2009.igem.org/User:DavidC/19_September_2009 19] [http://2009.igem.org/User:DavidC/20_September_2009 20]
[http://2009.igem.org/User:DavidC/21_September_2009 21] [http://2009.igem.org/User:DavidC/22_September_2009 22] [http://2009.igem.org/User:DavidC/23_September_2009 23] [http://2009.igem.org/User:DavidC/24_September_2009 24] [http://2009.igem.org/User:DavidC/25_September_2009 25] [http://2009.igem.org/User:DavidC/26_September_2009 26] [http://2009.igem.org/User:DavidC/27_September_2009 27]
[http://2009.igem.org/User:DavidC/28_September_2009 28] [http://2009.igem.org/User:DavidC/29_September_2009 29] [http://2009.igem.org/User:DavidC/30_September_2009 30]
October
MTWTFSS
      [http://2009.igem.org/User:DavidC/1_October_2009 1] [http://2009.igem.org/User:DavidC/2_October_2009 2] [http://2009.igem.org/User:DavidC/3_October_2009 3] [http://2009.igem.org/User:DavidC/4_October_2009 4]
[http://2009.igem.org/User:DavidC/5_October_2009 5] [http://2009.igem.org/User:DavidC/6_October_2009 6] [http://2009.igem.org/User:DavidC/7_October_2009 7] [http://2009.igem.org/User:DavidC/8_October_2009 8] [http://2009.igem.org/User:DavidC/9_October_2009 9] [http://2009.igem.org/User:DavidC/10_October_2009 10] [http://2009.igem.org/User:DavidC/11_October_2009 11]
[http://2009.igem.org/User:DavidC/12_October_2009 12] [http://2009.igem.org/User:DavidC/13_October_2009 13] [http://2009.igem.org/User:DavidC/14_October_2009 14] [http://2009.igem.org/User:DavidC/15_October_2009 15] [http://2009.igem.org/User:DavidC/16_October_2009 16] [http://2009.igem.org/User:DavidC/17_October_2009 17] [http://2009.igem.org/User:DavidC/18_October_2009 18]
[http://2009.igem.org/User:DavidC/19_October_2009 19] [http://2009.igem.org/User:DavidC/20_October_2009 20] [http://2009.igem.org/User:DavidC/21_October_2009 21] [http://2009.igem.org/User:DavidC/22_October_2009 22] [http://2009.igem.org/User:DavidC/23_October_2009 23] [http://2009.igem.org/User:DavidC/24_October_2009 24] [http://2009.igem.org/User:DavidC/25_October_2009 25]
[http://2009.igem.org/User:DavidC/26_October_2009 26] [http://2009.igem.org/User:DavidC/27_October_2009 27] [http://2009.igem.org/User:DavidC/28_October_2009 28] [http://2009.igem.org/User:DavidC/29_October_2009 29] [http://2009.igem.org/User:DavidC/30_October_2009 30] [http://2009.igem.org/User:DavidC/31_October_2009 31]


Contents

Sunday the 13th

Restriction digest

Ligation between BBa_C0012 and BBa_B0014:

BBa_B0014 (=2,41µg/µL) restriction digest by EcoRI and XbaI (3284bp):

DNA (10µg final)= 4,2µL
Buffer Eco R1 (NEB) = 5µL
H20 = 39,8µL
Eco R1 = 1µL
Incubation 1h at 37°C.

DNA purification with a nucleospin (macherey-nagel) .

DNA = 50µL
Buffer 2 (NEB) = 6µL
BSA (NEB) = 0,6µL
H20 = 2,4µL
Xba 1 = 1µL
Incubation 1h at 37°C.

C0012 (1,55µg/µL) restriction digest by EcoRI and SpeI (1128bp):

DNA (10µg final) = 6,4µL
Buffer Eco R1 (NEB) = 5µL
H20 = 35,6µL
BSA = 0,5µL
Eco R1 (NEB) = 1µL
Spe 1 (NEB) = 1µL
Incubation 1h at 37°C.

TAKARA Enzyme testing

Ligation between BBa_C0012 and BBa_B0014:

B0014 (= 2,405µg/µL) restriction digest with EcoRI and XbaI (3128bp):

ADN (10µg final) = 4,2µL
Buffer M (TAKARA) = 5µL
H20 = 38,8µL
Eco R1 = 1µL
Xba 1 = 1µL
Incubation 1h à 37°C.

DNA electrophoresis

85 Volt, 15 minutes.
105 Volt, 40 minutes.
Ladder fermentas 1 Kb.

Samples:

DNA purification

Kit Qiagen “gel extraction kit”, final volume = 50µL.

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