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Team:Valencia/Parts
From 2008.igem.org
(Difference between revisions)
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Total DNA was extracted from our yeast strains.<br> | Total DNA was extracted from our yeast strains.<br> | ||
UCP-1 was amplified by PCR using oligonucleotides matching the sequence and bearing the appropriate Biobrick prefix and suffix.<br> | UCP-1 was amplified by PCR using oligonucleotides matching the sequence and bearing the appropriate Biobrick prefix and suffix.<br> | ||
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Primers' sequences (EcoRI and PstI sites in bold):<br> | Primers' sequences (EcoRI and PstI sites in bold):<br> | ||
Forward: 5' '''gAATTC'''gCggCCgCTTCTAgATggTgAgTTCgACAACTTC 3';<br> | Forward: 5' '''gAATTC'''gCggCCgCTTCTAgATggTgAgTTCgACAACTTC 3';<br> | ||
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<ol>72º 10'</ol> | <ol>72º 10'</ol> | ||
</ol> | </ol> | ||
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| + | '''Results:'''<br> | ||
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| + | [[Image:Valencia_PCRresults.jpg]]<br> | ||
| + | Intense amplicons, of the expected size, (about 1 kb) were produced for UCP-1, 175-deleted and 76 deleted. <br> | ||
| + | Amplicons were digested (H buffer) with EcoRI y PstI.<br> | ||
| + | |||
| + | |||
</div> | </div> | ||
</div> | </div> | ||
Revision as of 18:32, 29 October 2008
Construction of Valencia Team Biobricks
Preparing inserts
Total DNA was extracted from our yeast strains.
UCP-1 was amplified by PCR using oligonucleotides matching the sequence and bearing the appropriate Biobrick prefix and suffix.
Primers' sequences (EcoRI and PstI sites in bold):
Forward: 5' gAATTCgCggCCgCTTCTAgATggTgAgTTCgACAACTTC 3';
Reverse: 5' TACTAgTAgCggCCgCTgCAgCTATgTggTgCAgTCCACTg 3'
PCR was conducted as follows:
- A first denaturation cycle
- 94º 3'
Followed by 30 amplification cycles:
- 94º 30
60º 1' 30
72º 1'
And a final extension step:
- 72º 10'
Results:
Intense amplicons, of the expected size, (about 1 kb) were produced for UCP-1, 175-deleted and 76 deleted.
Amplicons were digested (H buffer) with EcoRI y PstI.
