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Team:HokkaidoU Japan/Projects/asB0034/Method
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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.
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
