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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

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

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

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