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Friday 14th August

Lab Work

Quantification

by Pauline

• BBa_R0051 (2nd send)
• BBa_K1707011 #3
• BBa_K1707012 #2
• BBa_K1707033 #1 and #2

Mix:

• 2 µL plasmid
• 2 µL DNA Loading 6X
• 6 µL H2O

Agarose gel 1%, Migration 90V

Quantification, from left to right: 1. DNA Ladder, 2. BBa_K1707011#3 (08/12), 3. BBa_K1707011#3 (08/12), 4. BBa_K1707012#2 (08/12), 5. BBa_K1707012#2 (08/13), 6. BBa_K1707033#1, 7. BBa_K1707033#2, 8. BBa_R0051 (2^{nd send), 9. Empty, 10. Empty}

We can conclude:

• BBa_R0051 (2nd send): 13 µg/µL
• BBa_K1707011 #3: 40 µg/µL
• BBa_K1707012 #2: 35 µg/µL
• BBa_K1707033 #1 and #2: 20 µg/µL

New culture

by Pauline

• BBa_K1707035
• BBa_K1707036

We put 2 new clones in 5mL LB + 5µL Chloramphenicol

Plasmid extraction

by Pauline

• BBa_K1707031 #3, #4, #5 and #6

With Macherey-Nagel Extraction kit

Digestion

by Pauline

• BBa_K1707031 #3, #4, #5 and #6

Mix for each:

• BBa_K1707011 and BBa_K1707012
  • 0,5 µL XbaI
  • 0,5 µL PstI
  • 1 µL FastDigest Buffer 10x
  • 6 µL H2O
  • 2 µL plasmid

Incubation 37°C, 1H30

Ligation

by Pauline

• BBa_K1707023: BBa_K1707012 and BBa_J23101
  • 5,9 µL BBa_K1707012
  • 5 µL BBa_J23101
  • 2 µL Ligase
  • 1,5 µL Buffer
  • 0,6 µL H2O

• BBa_K1707022: BBa_K1707011 and BBa_R0051
  • 5,4 µL BBa_K1707011
  • 7,6 µL BBa_R0051
  • 2 µL Ligase
  • 2 µL Buffer
  • 3 µL H2O

• BBa_K1707034: BBa_K1707033 and BBa_J23101
  • 11 µL BBa_K1707033
  • 5 µL BBa_J23101
  • 2 µL Ligase
  • 2 µL Buffer

Incubation 3h, 4°C

Electrophoresis

by Pauline

Agarose gel, 1%, Migration 90V

Verification of gel purification, from left to right: 1. DNA Ladder, 2. BBa_K1707030#3, 3. BBa_K1707031#4, 4. BBa_K1707031#5, 5. BBa_K1707031#6, 6. Empty, 7. Empty, 8. Empty, 9. Empty, 10. Empty

Biobricks:

• Verification:
  • BBa_K1707031 #3, #4, #5 and #6

We can conclude that all clones aren't OK because the band viewed corresponding to the BBa_R0040 plasmid

Transformation

by Pauline

• BBa_K1707022
• BBa_K1707023
• BBa_K1707034

Soil experiment

by Coralie and Johan

Low budget experiment

by Coralie

Member present:

• Instructors: Claire
• Students: Coralie and Pauline

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