"
Team:Paris Bettencourt/Notebook/TB-ception/Thursday 12th September.html
From 2013.igem.org
(Difference between revisions)
| Line 4: | Line 4: | ||
<a href="https://2013.igem.org/Team:Paris_Bettencourt/Project/TB-ception" target="_blank" class="tbnotelogo TClogo"> ASDF </a> | <a href="https://2013.igem.org/Team:Paris_Bettencourt/Project/TB-ception" target="_blank" class="tbnotelogo TClogo"> ASDF </a> | ||
<div style="clear: both;"></div> | <div style="clear: both;"></div> | ||
| - | <h3>Thursday<sup>th</sup> September</h3> | + | <h3>Thursday 12<sup>th</sup> September</h3> |
<p><b><em> | <p><b><em> | ||
Revision as of 13:32, 25 September 2013
TB-ception
ASDFThursday 12th September
• linear bacbone PCR: Phusion protocol from the 10th of the September, new PCR program: 98.0 (30 s) - 98.0 (10 s), 58.0 (15 s), 72.0 (1:10) - 72.0 (3 min). This time the PCR worked fine - no unspecific binding; the band was 2 kb long.
• PCR for biobricking TDMH: we designed primers to add biobrick cutsites on our TDMH gene. Template - pET21b+TDMH miniprep (150 ng/µL), primers - TDMH F (gaattcgcggccgcttctagATGATTTCCCTCCGGAAGCC) and TDMH R (ctgcagcggccgctactagtaTCAGTGGTGGTGGTGGTGGT). Program: 98.0 (30 s) - 98.0 (10 s), 62.0 - 58.0 (30 s), 72.0 (30 s) - 72.0 (10 min). It worked, bend size - 700 bp.
• Gibson assembly for the new backbones: PCR products from the 10th of September were used (the same PCR reaction was done for the biobrick pSB2C3 backbone with all primers, so each PCR product for the Duet backbones was mixed with the corresponding pSB product. Protocol: 5 µL of each product, 10 µL of the Gibson master mix, 15 min on 50.0.
