Team:Hawaii/Notebook/2008-09-25

From 2008.igem.org

(Difference between revisions)
(marg)
(Drylab Work)
 
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::*oriT + J33207(de-phosphorylated) colony 1
::*oriT + J33207(de-phosphorylated) colony 1
::*5 additional colonies of oriTr transformation were PCR'ed
::*5 additional colonies of oriTr transformation were PCR'ed
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== Drylab Work ==
 
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===Name of Task===
 
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:<strong> name of person/people who performed the task</strong>
 
-
 
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:* Summary of task and what was done. Link to experiment for detailed notes if necessary.
 
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:* e.g. read through papers, worked on proposal, etc.
 
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===Construction of secretion device and redo of BB-pRL1383a (cont.)===
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:<strong> Grace</strong>
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[[Image:092508REdigest.jpg|right|thumb|300px|EtBr stained 2% agarose gels ran at 60V for 1.5 hours and 2 hours, respectively. Thirty microliters of RE digested products were loaded into each well.]]
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:* Ran RE digests from last night on gel
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::* nrsg8 and nrg4 did not cut. Implies insert wrong?
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:* Extracted bands from gel
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:* Ligated:
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::* 3A using pSB1A3:
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:::* nrg7 + tt
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:::* rgt1 + nir
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:::* rgt1 + plac
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:::* rgt2 + nir
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:::* rgt2 + plac
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:::* pSB1A3 (control)
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:* pRL1383a + J33207
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::* pRL1383a to self (control)
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:* Transformed into DH5&alpha; using 5 &mu;l ligation reaction; also transformed J33207 and pRL1383a plasmids as control)
= Discussion =
= Discussion =

Latest revision as of 23:51, 25 September 2008

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

Things we did today

Wetlab work

Verification of Ligation

Margaret
  • The plac/rbs+rep and oriTr(BB) transformations resulted in 0 colonies with the correct insert, so the remainder of the ligation reaction from 9/16 was digested with XbaI and run on a gel.

Sequencing & Colony PCR

Margaret
  • PCR
  • oriV colonies (3,4,6,8,9,10,11)
  • aadA(BB) colonies (4,5,7,8)
  • aadA(pRL) colonies (1,2)
  • oriT + J33207 (1,2,4)
  • oriT + J33207(de-phosphorylated) colony 1
  • 5 additional colonies of oriTr transformation were PCR'ed


Construction of secretion device and redo of BB-pRL1383a (cont.)

Grace
EtBr stained 2% agarose gels ran at 60V for 1.5 hours and 2 hours, respectively. Thirty microliters of RE digested products were loaded into each well.
  • Ran RE digests from last night on gel
  • nrsg8 and nrg4 did not cut. Implies insert wrong?
  • Extracted bands from gel
  • Ligated:
  • 3A using pSB1A3:
  • nrg7 + tt
  • rgt1 + nir
  • rgt1 + plac
  • rgt2 + nir
  • rgt2 + plac
  • pSB1A3 (control)
  • pRL1383a + J33207
  • pRL1383a to self (control)
  • Transformed into DH5α using 5 μl ligation reaction; also transformed J33207 and pRL1383a plasmids as control)

Discussion

Quote of the Day

History is the only laboratory we have in which to test the consequences of thought. - Étienne Gilson


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