Team:UNICAMP-Brazil/Notebooks/September 25

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''Marcelo''
''Marcelo''
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==== PY Promoter - Colony-PCR screening ====
 
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*<p style=”text-align:justify;”>We selected 5 white colonies of each transformation we did yesterday to perform a colony-PCR screening. In this PCR we used the M13 primers of pGEM vector. In those colonies that have the vector with our fragment inserted this pair of primers would amplify a fragment with XX bp (PY1) and XX bp (PY2).</p>
 
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*<p style=”text-align:justify;”>We analyzed the results of our PCR in an agarose gel:</p>
 
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[[Image:py_gel_3.png|400px|center]]
 
==== CeiB: Searching the right colony ====
==== CeiB: Searching the right colony ====
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*<p style=”text-align:justify;”>We decided to search in the plate possible cells with the right DNA insert. We made 15 colony of each plate.  Unfortunately we didn’t get again.
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*<p style=”text-align:justify;”>We decided to search in the plate possible cells with the right DNA insert. We made 15 colony of each plate.  Unfortunately we didn’t get it again.</p>
''Ane''
''Ane''
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====Biofusion vector - Electroelution====
====Biofusion vector - Electroelution====
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*<p style=”text-align:justify;”> Considering that we need more vector to perform the ligation reactions, we electroeluted more vector from  the ADH1 biobrick previously digested with ''Xba''I and ''Spe''I ([https://2009.igem.org/Team:UNICAMP-Brazil/Protocols/Electroelution Protocol 12]).</p>
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*<p style=”text-align:justify;”>Considering that we need more vector to perform the ligation reactions, we electroeluted more vector from  the ''ADH1'' biobrick previously digested with ''Xba''I and ''Spe''I ([https://2009.igem.org/Team:UNICAMP-Brazil/Protocols/Electroelution Protocol 12]).</p>
''Taís''
''Taís''
{{:Team:UNICAMP-Brazil/inc_rodape}}
{{:Team:UNICAMP-Brazil/inc_rodape}}

Latest revision as of 03:44, 22 October 2009

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ColiGuard

Getting the visa

  • Today was a happy day to five of us! We went to the U.S. Embassy in Sao Paulo and we got our visas! :D

finO and finP - Still Trying to Confirm our Biobricks

  • We ran an agarose gel of yesterday's PCRs product.

  • We couldn't obtain even a single amplified fragment! =(

  • Why can the transformed cells grew in the media (that means they actually have the AMP resistence), but they haven't our inserts into it's plasmids?

  • Our advisors suggested us that, since we digested our plasmid vector with XbaI and SpeI (X sticky ends can come together with S sticky ends), it recircularized without the introduction of our inserts.

  • Therefore, we must perform the dephosphorylation of our digested vector. That may prevent it from recircularizing without the insert introduction.

Marcelo


CeiB: Searching the right colony

  • We decided to search in the plate possible cells with the right DNA insert. We made 15 colony of each plate. Unfortunately we didn’t get it again.

Ane

YeastGuard

Biofusion vector - Electroelution

  • Considering that we need more vector to perform the ligation reactions, we electroeluted more vector from the ADH1 biobrick previously digested with XbaI and SpeI (Protocol 12).

Taís




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