Team:Hawaii/Notebook/2008-08-30

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(Difference between revisions)
(New page: {{Team:Hawaii/Header}} = Things we did today = == Wetlab work == ===Construction of p+r and rereplacement of BB-pRL1383a MCS=== :<strong> Grace</strong> [[Image:083008REdigests.jpg|right|...)
(→Construction of p+r and rereplacement of BB-pRL1383a MCS)
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[[Image:083008REdigests.jpg|right|thumb|400px|EtBr stained 2% (L) and 0.8% (R) agarose gels ran at 60V for 2 hours and 1 hour, respectively. Thirty microliters of RE digest reaction were loaded into each well.]]
[[Image:083008REdigests.jpg|right|thumb|400px|EtBr stained 2% (L) and 0.8% (R) agarose gels ran at 60V for 2 hours and 1 hour, respectively. Thirty microliters of RE digest reaction were loaded into each well.]]
:* Ran RE digests from last night on agarose gel
:* Ran RE digests from last night on agarose gel
 +
::* C0012: correct vector band ~2.1kb
 +
::* J33207: band at ~700bp (only one enzyme cut -- XbaI did not cut; PstI cuts because C0012 was cut by PstI) and ~850bp (uncut PCR product)
 +
::* nir: ~800bp band (from GFP contaminant in plasmid prep, cut only once) and ~2kb band (??)
 +
::* BB-pRL1383a: ran WAY too much product. Will redo RE digest with 20% of plasmid used. Since J33207 was only cut once, it's assumed BB-pRL1383a was cut only once as well.
:* Extracted bands from gel
:* Extracted bands from gel
:* Ligated:
:* Ligated:
-
::* BB-pRL1383a with J33207
 
-
::* nir + rbs (B0034)
 
::* plac + rbs (B0034)
::* plac + rbs (B0034)
-
:* RE digested p+r with EcoRI, XbaI, SpeI, PstI
+
:* PCR of J33207
 +
:* RE digested:
 +
::* p+r with EcoRI, XbaI, SpeI, PstI
 +
::* J33207 and BB-pRL1383a with EcoRI and PstI
 +
:* Treated C0012 vector (pSB1A2) with SAP
= Discussion =
= Discussion =

Revision as of 23:52, 30 August 2008

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Things we did today

Wetlab work

Construction of p+r and rereplacement of BB-pRL1383a MCS

Grace
EtBr stained 2% (L) and 0.8% (R) agarose gels ran at 60V for 2 hours and 1 hour, respectively. Thirty microliters of RE digest reaction were loaded into each well.
  • Ran RE digests from last night on agarose gel
  • C0012: correct vector band ~2.1kb
  • J33207: band at ~700bp (only one enzyme cut -- XbaI did not cut; PstI cuts because C0012 was cut by PstI) and ~850bp (uncut PCR product)
  • nir: ~800bp band (from GFP contaminant in plasmid prep, cut only once) and ~2kb band (??)
  • BB-pRL1383a: ran WAY too much product. Will redo RE digest with 20% of plasmid used. Since J33207 was only cut once, it's assumed BB-pRL1383a was cut only once as well.
  • Extracted bands from gel
  • Ligated:
  • plac + rbs (B0034)
  • PCR of J33207
  • RE digested:
  • p+r with EcoRI, XbaI, SpeI, PstI
  • J33207 and BB-pRL1383a with EcoRI and PstI
  • Treated C0012 vector (pSB1A2) with SAP

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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