IGEM:Harvard/2006/Cyanobacteria/Notebook/2006-7-21

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Contents

To-do

  1. Inoculate liquid cultures with retransformed TP strain from yesterday-- done
  2. Extract and purify LC1PUR1 and LC2PUR2 (3kb and 400b bands) from PCR Extraction #5 of 2006-7-19. --done
  3. Ligate gel-purified PCR product with Topo, transform with Top10 competent cells, spread plates
  4. Wiki cleanup-- in progress
  5. Update incubator status
  6. Work on Monday presentation outline
  7. Send out signed synthesis contract with GeneArt-- done

Strike three

Once again we left a bucket of ice out on the benchtop overnight. This one contained Invitrogen's Zero-Blunt Topo cloning kit. We've labeled the box as possibly bad.

Synthesis contract

  1. Sign contract (3x)
  2. Billing - mailto address needs to be changed
  3. Add comment to ship as soon as ready

Update: Alain signed the analyses pages on behalf of Peng. We also corrected the shipping and billing addresses on the last page of the price quotation. We faxed all four pages to Stefan Weinges at Geneart around 3:15PM.

Inoculation

  • Each of the plates that were transformed from last night had lots of growth.
  • One inoculation was made for each of the plates.
  • One colony was taken from each of the plates to prevent mixes of colonies being grown up.
  • The inoculations consisted of:
    • 5 mL LB liquid culture
    • 2 um Kan
    • one colony

Inoculated around noon.

Gel extraction

We extracted 4 different bands from the gel that we ran yesterday:

  • LC1PUR1 3kb fragment
    • 85mg
  • LC1PUR1 400bp fragment
    • 197.5mg
  • LC2PUR1 3kb fragment
    • 132.9mg
  • LC2PUR1 400bp fragment
    • 213.3mg

The remainder of the gel has been labeled and placed in a refrigerator, in case we're interested in the other lanes.

Gel Purification

We purified the gels using the QiaQuick Gel Extraction Kit. DNA was eluted with 30 ┬ÁL H2O.

  • LC1PUR1 3kb fragment
    • Use 255 ul of Buffer QG
  • LC1PUR1 400bp fragment
    • 592.5 ul
  • LC2PUR1 3kb fragment
    • 398.7 ul
  • LC2PUR1 400bp fragment
    • 639.9 ul
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