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Revision as of 10:28, 28 September 2007 (view source) Blent (Talk | contribs) (→pOmpC glucose response testing by construct (pOmpC + EYFP)) ← Older edit		Revision as of 10:30, 28 September 2007 (view source) Blent (Talk | contribs) (→pOmpC glucose response testing by construct (pOmpC + EYFP)) Newer edit →
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Revision as of 10:30, 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]

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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