Team:HokkaidoU Japan/Projects/asB0034/Method

From 2014.igem.org

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<h1>How to synthesize antisense RNA
<h1>How to synthesize antisense RNA
<h4><p>Antisense-RBS fragments were synthesized based on BioBrick by primer annealing. The sides of antisense fragment have scar and restriction enzymes XhoI, NcoI.</p>
<h4><p>Antisense-RBS fragments were synthesized based on BioBrick by primer annealing. The sides of antisense fragment have scar and restriction enzymes XhoI, NcoI.</p>
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<div class="fig fig400">
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<img src="http://hoge">
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<div>caption</div>
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</div>
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<div class="fig fig400 para">
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<img src="https://static.igem.org/mediawiki/2014/0/02/HokkaidoU_antisenseB0034_overview07.png">
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</div>
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<div>How to make antisense B0034 by primer annealing</div>
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</div>
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<div class="fig fig400 para">
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<img src="https://static.igem.org/mediawiki/2014/6/63/HokkaidoU_antisenseB0034_overview08.png">
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</div>
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<div>Using restriction enzyme, XhoI, NcoI, we made stem-antisense conplex. </div>
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</div>
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<div class="fig fig400">
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<img src="https://static.igem.org/mediawiki/2014/c/c1/HokkaidoU_antisenseB0034_overview06.png">
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<div>Primer for synthesizing antisense B0034</div>
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</div>
<h1>How to assay
<h1>How to assay
<h4><p>we selected RFP and GFP for target genes. We used fluorophotometer to measure how antisense worked. The colonies transformed antisense RNA and target gene used to do assay.</p>
<h4><p>we selected RFP and GFP for target genes. We used fluorophotometer to measure how antisense worked. The colonies transformed antisense RNA and target gene used to do assay.</p>
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<p>(1)To cultivate the colony in 2 mL LB culture for 9 hours</p>
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<p>(1)To cultivate the colony in 4 mL LB culture for about 20 hours</p>
<p>(2)To control turbidity up to 0.1 at OD600</p>
<p>(2)To control turbidity up to 0.1 at OD600</p>
<p>(3)To cultivate the colony in 2 mL M9ZB culture for 9 hours (IPTG induces antisense RNA, addition 20 uL )</p>
<p>(3)To cultivate the colony in 2 mL M9ZB culture for 9 hours (IPTG induces antisense RNA, addition 20 uL )</p>
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<p>(4)To measure fluorescence after 9hours, 11hours, 13hours </p>
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<p>(4)To measure fluorescence after 9 hour </p>
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<div class="fig fig800">
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<img src="https://static.igem.org/mediawiki/2014/2/25/HokkaidoU_antisenseB0034_overview09.png">
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<div>Antisense B0034 is induce by IPTG</div>
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</div>
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Revision as of 07:37, 14 October 2014

How to synthesize antisense RNA

Antisense-RBS fragments were synthesized based on BioBrick by primer annealing. The sides of antisense fragment have scar and restriction enzymes XhoI, NcoI.

caption
How to make antisense B0034 by primer annealing

Using restriction enzyme, XhoI, NcoI, we made stem-antisense conplex.
Primer for synthesizing antisense B0034

How to assay

we selected RFP and GFP for target genes. We used fluorophotometer to measure how antisense worked. The colonies transformed antisense 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

Antisense B0034 is induce by IPTG

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