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Team:HokkaidoU Japan/Projects/asB0034/Method
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<div id="hokkaidou-contents"> | <div id="hokkaidou-contents"> | ||
| - | <h1>How to synthesize | + | <h1>How to synthesize anti-sense RNA |
| - | <h4><p> | + | <h4><p>Anti-sense RBS fragments were synthesized based on BioBrick by primer annealing. The sides of antisense fragment have scar and restriction enzymes XhoI, NcoI.</p> |
<div class="fig fig400 para"> | <div class="fig fig400 para"> | ||
<img src="https://static.igem.org/mediawiki/2014/0/02/HokkaidoU_antisenseB0034_overview07.png"> | <img src="https://static.igem.org/mediawiki/2014/0/02/HokkaidoU_antisenseB0034_overview07.png"> | ||
| - | <div>Fig1. How to make | + | <div>Fig1. How to make anti-sense B0034 by primer annealing</div> |
</div> | </div> | ||
<div class="fig fig400 para"><p> | <div class="fig fig400 para"><p> | ||
<img src="https://static.igem.org/mediawiki/2014/6/63/HokkaidoU_antisenseB0034_overview08.png"> | <img src="https://static.igem.org/mediawiki/2014/6/63/HokkaidoU_antisenseB0034_overview08.png"> | ||
| - | <div>Fig2. Using restriction enzyme, XhoI, NcoI, we made | + | <div>Fig2. Using restriction enzyme, XhoI, NcoI, we made stem_anti-sense conplex. </div> |
</div> | </div> | ||
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<h1><p>How to assay</p> | <h1><p>How to assay</p> | ||
| - | <h4><p>we selected RFP and GFP for target genes. We used fluorophotometer to measure how | + | <h4><p>we selected RFP and GFP for target genes. We used fluorophotometer to measure how anti-sense worked. The colonies transformed anti-sense RNA and target gene used to do assay.</p> |
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<div class="fig fig800"> | <div class="fig fig800"> | ||
<img src="https://static.igem.org/mediawiki/2014/2/25/HokkaidoU_antisenseB0034_overview09.png"> | <img src="https://static.igem.org/mediawiki/2014/2/25/HokkaidoU_antisenseB0034_overview09.png"> | ||
| - | <div>Fig4. | + | <div>Fig4. Anti-sense B0034 is induced by IPTG</div> |
</div> | </div> | ||
Revision as of 08:58, 14 October 2014
How to synthesize anti-sense RNA
Anti-sense RBS fragments were synthesized based on BioBrick by primer annealing. The sides of antisense fragment have scar and restriction enzymes XhoI, NcoI.
Fig1. How to make anti-sense B0034 by primer annealing
Fig2. Using restriction enzyme, XhoI, NcoI, we made stem_anti-sense conplex.
Fig3. Blue; antisense B0034, B0032 Red; scar sequence Green; NcoI site Purple; XhoI site
How to assay
we selected RFP and GFP for target genes. We used fluorophotometer to measure how anti-sense worked. The colonies transformed anti-sense RNA and target gene used to do assay.
(1)To cultivate the colony in 4 mL LB culture for about 20 hours
(2)To control turbidity up to 0.1 at OD600
(3)To cultivate the colony in 2 mL M9ZB culture for 9 hours (IPTG induces antisense RNA, addition 20 uL )
(4)To measure fluorescence after 9 hour
Fig4. Anti-sense B0034 is induced by IPTG
