Team:UNICAMP-Brazil/Notebooks/October 9
From 2009.igem.org
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* We decided that we are going to confirm such ligations by performing: | * We decided that we are going to confirm such ligations by performing: | ||
- | -A Digestion: with | + | -A Digestion: with ''Xba''I and ''Spe''I restriction enzymes, in order to release our parts from pGEM plasmid; |
-A PCR: performed with the specific forward primer for our insert and with the reverse primer for pGEM plasmid (M13), in order to confirm that, once our inserts are indeed in there, they are also in the correct frame position. | -A PCR: performed with the specific forward primer for our insert and with the reverse primer for pGEM plasmid (M13), in order to confirm that, once our inserts are indeed in there, they are also in the correct frame position. | ||
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''Fabi and Léo'' | ''Fabi and Léo'' | ||
- | ====Cre-Recombinase - Confirmation: | + | ====Cre-Recombinase - Confirmation: Stage I==== |
*<p style=”text-align:justify;”>After performing minipreps (on October 5th) for 20 inoculated cultures, possibly harbouring our Cre-Recombinase without ATG's Biobricks, it's time to start the confirmation procudures. Regarding "confirmation procudures", we mean a two stages procedure: (1) perform a specific PCR for Cre-Recombinase and (2) perform a digestion, aiming in excising a fragment of expectable size from the pSB1A3 vector.</p> | *<p style=”text-align:justify;”>After performing minipreps (on October 5th) for 20 inoculated cultures, possibly harbouring our Cre-Recombinase without ATG's Biobricks, it's time to start the confirmation procudures. Regarding "confirmation procudures", we mean a two stages procedure: (1) perform a specific PCR for Cre-Recombinase and (2) perform a digestion, aiming in excising a fragment of expectable size from the pSB1A3 vector.</p> | ||
*<p style=”text-align:justify;”>Today we did stage I, a PCR reaction using Cre's specific designed primers, running an agarose gel straight after reaction ended.</p> | *<p style=”text-align:justify;”>Today we did stage I, a PCR reaction using Cre's specific designed primers, running an agarose gel straight after reaction ended.</p> | ||
*<p style=”text-align:justify;”>Thereby, according to Stage I.... we did it! Several samples appears to contain our Cre's Biobrick!</p> | *<p style=”text-align:justify;”>Thereby, according to Stage I.... we did it! Several samples appears to contain our Cre's Biobrick!</p> | ||
- | *<p style=”text-align:justify;”>If we succeed in Stage II, we will finally be able to say we got a biobrick! =] | + | *<p style=”text-align:justify;”>If we succeed in Stage II, we will finally be able to say we got a biobrick! =]</p> |
''Víctor'' | ''Víctor'' | ||
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==''' YeastGuard '''== | ==''' YeastGuard '''== | ||
====[https://2009.igem.org/Team:UNICAMP-Brazil/Protocols/pGEMStrategy New Strategy: pGEM]==== | ====[https://2009.igem.org/Team:UNICAMP-Brazil/Protocols/pGEMStrategy New Strategy: pGEM]==== | ||
- | *<p style= | + | *<p style="text-align:justify;">The plates containing ''E. coli'' transformed with pDLD didn’t grow. We repeated the ligation reaction and transformed competent ''E. coli'' again.</p> |
*<p style=”text-align:justify;”>We transformed the ligation between Lysozyme and biofusion, performed yesterday, in competent ''E. coli'' and plated in LB+Amp media.</p> | *<p style=”text-align:justify;”>We transformed the ligation between Lysozyme and biofusion, performed yesterday, in competent ''E. coli'' and plated in LB+Amp media.</p> |
Latest revision as of 03:20, 22 October 2009
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