NYMU Taipei/Lab Notes/2007 9 27

(Difference between revisions)

Revision as of 10:22, 28 September 2007

CinR+HSL+D-term check
- After re-culture in 20-block plate,randomly pick 6 blocks from CinR+HSL+D-term plate
- PCR check using VF2, VR primers
- PCR total 50 uL, 10 uL for GEL separation
- sample at each lane
  - 1Kb ladder
  - block #1 (insert band appears)
  - block #2 (insert band appears, and one extra band around 400 bp)
  - block #3 (insert band appears)
  - block #4 (insert band appears)
  - block #5 (insert band does not appear, only band around 400 bp)
  - block #6 (insert band does not appear, only band around 400 bp)
- expect results: one band at 1,623 bp is our insert
  - insert = RBS+CinR (2,843 - 2,053) + RBS+HSL (2,783 - 2,053) + D-term (3,266 - 3,163)
  - insert = 790 + 730 + 103 = 1,623
- extra band at 400 bp might be plasmid failed to be correctly ligated
  - PCR product length without any insert = (3,189 - 3,049) + 176 = 316
- References
  - [http://partsregistry.org/Image:PsB1AK3_map.jpg vector map of pSB1AK3 and VF2 VR primers]

pOmpC glucose response testing by construct (pOmpC + EYFP)
- design: 5 tubes for 0%, 1%, 3%, 5%, 10% extra glucose cultured in LB medium
- tube with 0% extra glucose is treated as negative control

glucose (%)	O.D. 600	white light	green light
0	2.487
1	2.091
3	1.861
5	1.637
10	1.025

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