Registry/Measurement kit/Notebook/2007-6-15

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Contents

Plan for today

  1. Digest constructs
  2. Ligate components
  3. Transform

Double Digest

  • See protocol
  • Used buffer 3 as suggested by NEB for the XbaI/PstI digests
  • Used 2uL stock R0040 instead of prepped
  • 22.12 uL of B0032
  • 8.84 uL of I13401
  • 4.68 uL of J04650
  • Incubated at 37 for 5 hrs

Design discussion

See proposed solution at Registry/Measurement_kit/Notebook/Design

  • It was suggested that we use the LacZ reporter gene instead of an FP, but all that appears to be in the registry is an N-terminus fragment of the LacZ gene. In the near future, we plan to standardize and enter the LacZ gene into the registry.

PCR purification

  • buffer transfer of cut DNA
  • protocol is in booklet with the kit
  • note: labeling system for digests:
  1. R0040
  2. B0032
  3. I13401
  4. J04650

Ligation

Endy:DNA_ligation_using_T4_DNA_ligase

  • Used modified version of above protocol. Will post on OWW soon.

Labels: A: RFP promoter tester

  • B0032 + J04650

B: GFP RBS tester

  • R0040 + I13401

C: RFP RBS tester

  • R0040 + J0465

Transformation of devices

  • Attempting to transform the ligated devices.
  • Strain - MG1655
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