Team:Paris Saclay/Protocols/BioBrick Assembly


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BioBrick Assembly

This protocol is based on the original 3A Assembly from iGEM. We modified it to a assembly that places the core of one BioBrick - here called Part A - into another BioBrick - here called Part B.

TODO: Process illustration [Format I and Format II]



Note: Manipulate the enzymes with a proper subzero temperature support.

Part A:

  • Enzyme A: EcoRI
  • Enzyme B: SpeI

Part B with Format I (Part B placed after Part A):

  • Enzyme A: EcoRI
  • Enzyme B: XbaI

Part B with Format II (Part B placed before Part A):

  • Enzyme A: SpeI
  • Enzyme B: PstI

Digest reaction

  1. Take a 1.5ml microcentrifuge tube (Eppendorf) and put:
    1. xμl of H20 milliQ (complement)
    2. 5μl of buffer (Fast Digest Buffer 10x)
    3. xμl of Part A DNA
    4. 1μl of Enzyme A
    5. 1μl of Enzyme B
  2. Repeat step 1 with Part B
  3. Mix gently both tubes
  4. Incubate at 37°C for one hour
  5. Store Part B at -20°C.

Segregate process

  1. part A
  2. big box, gel 0.8% agarose (and BET concentration???????)
  3. 1 hour
  4. minimal exposure of UV light
  5. mass of the ependorf (before / after)

DNA extration from agarose gels