Difference between revisions of "Team:Paris Saclay/Notebook/August/12"

(Created page with "=Wednesday 12th August= ==Lab Work== ===Purification=== ''by Pauline'' Biobricks: * BBa_I13602 #1 * BBa_I13602 #2 * BBa_K1707020 * BBa_K1707030 With Macherey-Nagel Purificatio...")
 
m
 
(2 intermediate revisions by 2 users not shown)
Line 1: Line 1:
 +
{{Team:Paris_Saclay/notebook_header}}
 
=Wednesday 12th August=
 
=Wednesday 12th August=
 
==Lab Work==
 
==Lab Work==
Line 5: Line 6:
 
''by Pauline''
 
''by Pauline''
  
−
Biobricks:
 
 
* BBa_I13602 #1
 
* BBa_I13602 #1
 
* BBa_I13602 #2
 
* BBa_I13602 #2
Line 12: Line 12:
  
 
With Macherey-Nagel Purification kit
 
With Macherey-Nagel Purification kit
−
 
  
 
===Quantification===
 
===Quantification===
Line 18: Line 17:
  
 
Agarose gel 1%, Migration 90V
 
Agarose gel 1%, Migration 90V
−
 
+
[[File:ParisSaclay 12.08.15-quantif.jpg|300px|center]]
 +
<html><p><i>Quantification, from left to right: 1. <a href="https://2015.igem.org/File:Paris_Saclay-Ladder.jpg" target="_blank">DNA Ladder</a>, 2. BBa_K1707020, 3. BBa_K1707030, 4. BBa_I13602#1, 5. BBa_I13602#2,  6. Empty, 7. Empty, 8. Empty, 9. Empty, 10. Empty</i></p></html>
 
We can conclude:
 
We can conclude:
 
* BBa_I13602 #1: 40 µg/µL
 
* BBa_I13602 #1: 40 µg/µL
Line 24: Line 24:
 
* BBa_K1707020: 10 µg/µL
 
* BBa_K1707020: 10 µg/µL
 
* BBa_K1707030: 13 µg/µL
 
* BBa_K1707030: 13 µg/µL
−
 
−
 
  
 
===Digestion===
 
===Digestion===
 
''by Coralie''
 
''by Coralie''
  
−
Biobricks:
 
 
* BBa_K1707011 #3 and BBa_K1707012 #2
 
* BBa_K1707011 #3 and BBa_K1707012 #2
 
** Mix for each reaction:
 
** Mix for each reaction:
Line 39: Line 36:
 
*** 10 µL plasmid
 
*** 10 µL plasmid
 
** Incubation 37°C, 2h
 
** Incubation 37°C, 2h
−
 
  
 
===Purification on gel===
 
===Purification on gel===
Line 46: Line 42:
 
Agarose gel 1%, 90V
 
Agarose gel 1%, 90V
  
−
Biobricks:
 
 
* BBa_K1707011 #3
 
* BBa_K1707011 #3
 
* BBa_K1707012 #2
 
* BBa_K1707012 #2
 +
[[File:ParisSaclay 12.08.15-vérif purif.jpg|300px|center]]
 +
<html><p><i>Verification of gel purification, from left to right: 1. <a href="https://2015.igem.org/File:Paris_Saclay-Ladder.jpg" target="_blank">DNA Ladder</a>, 2. BBa_K1707011#3, 3. BBa_K1707012#2, 4. Empty, 5. Empty,  6. Empty, 7. Empty, 8. Empty, 9. Empty, 10. Empty</i></p></html>
  
 
We can conclude that biobricks are OK
 
We can conclude that biobricks are OK
  
 
Purification by the Macherey-Nagel Purification kit
 
Purification by the Macherey-Nagel Purification kit
−
 
−
 
  
 
===Ligation===
 
===Ligation===
Line 72: Line 67:
 
** 2,5 µL Buffer 10x
 
** 2,5 µL Buffer 10x
 
** 3,75 µL H2O
 
** 3,75 µL H2O
−
 
  
 
Incubation 4°C, ON
 
Incubation 4°C, ON
−
 
  
 
===New culture===
 
===New culture===
 
''by Pauline''
 
''by Pauline''
  
−
Biobricks:
 
 
* From stock:
 
* From stock:
 
** BBa_K1707012 #2 x2
 
** BBa_K1707012 #2 x2
Line 92: Line 84:
 
In 5mL LB + 5µL of appropriate antibiotic
 
In 5mL LB + 5µL of appropriate antibiotic
  
−
 
+
===Soil experiment===
−
 
+
  
 
'''Member present:'''
 
'''Member present:'''

Latest revision as of 20:15, 18 September 2015

Wednesday 12th August

Lab Work

Purification

by Pauline

  • BBa_I13602 #1
  • BBa_I13602 #2
  • BBa_K1707020
  • BBa_K1707030

With Macherey-Nagel Purification kit

Quantification

by Pauline

Agarose gel 1%, Migration 90V

ParisSaclay 12.08.15-quantif.jpg

Quantification, from left to right: 1. DNA Ladder, 2. BBa_K1707020, 3. BBa_K1707030, 4. BBa_I13602#1, 5. BBa_I13602#2, 6. Empty, 7. Empty, 8. Empty, 9. Empty, 10. Empty

We can conclude:

  • BBa_I13602 #1: 40 µg/µL
  • BBa_I13602 #2: 40 µg/µL
  • BBa_K1707020: 10 µg/µL
  • BBa_K1707030: 13 µg/µL

Digestion

by Coralie

  • BBa_K1707011 #3 and BBa_K1707012 #2
    • Mix for each reaction:
      • 1 µL PstI
      • 1 µL XbaI
      • 2 µL Buffer FastDigest 10x
      • 6 µL H2O
      • 10 µL plasmid
    • Incubation 37°C, 2h

Purification on gel

by Pauline

Agarose gel 1%, 90V

  • BBa_K1707011 #3
  • BBa_K1707012 #2
ParisSaclay 12.08.15-vérif purif.jpg

Verification of gel purification, from left to right: 1. DNA Ladder, 2. BBa_K1707011#3, 3. BBa_K1707012#2, 4. Empty, 5. Empty, 6. Empty, 7. Empty, 8. Empty, 9. Empty, 10. Empty

We can conclude that biobricks are OK

Purification by the Macherey-Nagel Purification kit

Ligation

by Pauline

  • BBa_K1707035: BBa_K1707030 + BBa_I13602
    • 11,6 µL BBa_K1707030
    • 1,25 µL BBa_R0040
    • 2 µL Ligase
    • 2 µL Buffer 10x
    • 3,15 µL H2O
  • BBa_K1707036: BBa_K1707020 + BBa_I13602
    • 15,5 µL BBa_K1707006
    • 1,25 µL BBa_B0030
    • 2 µL Ligase
    • 2,5 µL Buffer 10x
    • 3,75 µL H2O

Incubation 4°C, ON

New culture

by Pauline

  • From stock:
    • BBa_K1707012 #2 x2
    • BBa_K1707011 #3 x2
    • BBa_R0051
  • Two clones from plate of each:
    • BBa_K1707033
    • BBa_K1707031
    • BBa_K1707016

In 5mL LB + 5µL of appropriate antibiotic

Soil experiment

Member present:

  • Instructors: Claire
  • Students: Coralie and Pauline

Back to the calendar

Contact
eigem.parissaclay AT gmail DOT com
t@iGEMParisSaclay
aUniversité Paris Sud
a91405 Orsay, France Copyright © 2015 iGEM Paris Saclay. All rights reserved All inspirations encouraged.
*This wiki was conceived during long nights of work and a great coffee experience in the beautiful Orsay, France.