Team:Hawaii/Notebook/2008-08- 7

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::* Looks like nir+rbs and I14032+rbs worked. YAY :o). Will prep for sequencing tomorrow.
::* Looks like nir+rbs and I14032+rbs worked. YAY :o). Will prep for sequencing tomorrow.
::* GFP, GFPf, J33207 don't look like they went in. Will sequence to confirm anyway.
::* GFP, GFPf, J33207 don't look like they went in. Will sequence to confirm anyway.
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===Plasmid Prep===
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<strong>Margaret</strong>
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[[Image:Plasmid_prep_psmc121.jpg|right|thumb|300px|Plasmid Prep of pSMC121.]]
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:*Re-did yesterday's plasmid prep of pSMC121 because I don't believe the band was big enough. I e-mailed Dr. Callahan to get the exact size, but I know, because it contains the omega interposon, that it is at least 2kb.
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===Extraction from Gel===
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<strong>Margaret</strong>
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:*extracted 3 bands of 50ul reaction for rep and oriV first by cutting out using long wave UV for visualization.
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:* Purification from gel with Qiagen Minelute gel purification kit.
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===PCR===
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<strong>Margaret</strong>
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:*Large-scale PCR amplification of  aadA (it was lost in a gel cutting mishap) and p1 lytic.
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= Discussion =
= Discussion =

Revision as of 19:13, 8 August 2008

Projects Events Resources
Sponsors Experiments Milestones Protocols
Notebook (t) Meetings (t)

Things we did today

Wetlab work

Plasmid prep

Krystle
  • Finished plasmid prep of BB-pRL1383a, nir, GFPf from yesterday

Cyanobacteria seed stocks

Krystle
  • Reinoculated liquid and solid media seed stocks

Verification of transformants

EtBr stained 2.5% agarose gel ran at 95V for 75 min. Ten microliters of the PCR reaction were loaded into each well.
Grace
Construct Colony forming units
nir + B0030 (rbs) 2 + 1 cluster of colonies
plac (I14032) + B0030 (rbs) 37
GFP (E0040) + B0015 (tt) 8
GFPf + B0015 (tt) 4
lac device (J33207) + B0015 (tt) 56
  • Colony PCR of transformants
  • Ran on EtBr stained 2.5% agarose gel at 95V for 75 min.
  • Looks like nir+rbs and I14032+rbs worked. YAY :o). Will prep for sequencing tomorrow.
  • GFP, GFPf, J33207 don't look like they went in. Will sequence to confirm anyway.


Plasmid Prep

Margaret

Plasmid Prep of pSMC121.
  • Re-did yesterday's plasmid prep of pSMC121 because I don't believe the band was big enough. I e-mailed Dr. Callahan to get the exact size, but I know, because it contains the omega interposon, that it is at least 2kb.

Extraction from Gel

Margaret

  • extracted 3 bands of 50ul reaction for rep and oriV first by cutting out using long wave UV for visualization.
  • Purification from gel with Qiagen Minelute gel purification kit.

PCR

Margaret

  • Large-scale PCR amplification of aadA (it was lost in a gel cutting mishap) and p1 lytic.


Discussion

Quote of the Day

Gernot: "So would you eat the E. coli to save them if we lose the -80C in like a war?"
Loren: "Well, I'll eat them, but you'll be collecting them."


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