Team:Grenoble/Biology/Protocols/AND test
From 2012.igem.org
(Difference between revisions)
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<section> | <section> | ||
<h1>"AND" gate test</h1> control of pBAD expression by cAMP and arabinose. | <h1>"AND" gate test</h1> control of pBAD expression by cAMP and arabinose. | ||
- | + | <h2>Goal</h2> | |
+ | Measure the dependence of the activity of the pBAD promoter on the concentrations of cAMP and arabinose. | ||
</section> | </section> | ||
- | + | <section> | |
- | + | <h2>Procedure</h2> | |
+ | <h3>Outline</h3> | ||
+ | The day before: Grow BW25113 cya<span class="exposant">-</span> containing the pAra/Bad_RBS_GFP plasmid of Alon in LB + Kanamycin. Leave on the shaker at 37°C until the next day morning. | ||
+ | Measure expression in different concentrations of arabinose and cAMP in M9 added with glucose, acetate or glycerol. | ||
</section> | </section> | ||
+ | <section> | ||
+ | <h2>Protocol</h2> | ||
+ | <h3>Outline</h3> | ||
+ | Add 10 microliters of an appropriate mix of cAMP and arabinose to each well of a 96-well microplate (see plate layout below). Add 130 microliters of M9 into each well. The medium is prepared fresh, but not filtered. Then, add 10 microliters of the o/n culture and read in Fusion at 37°C. | ||
+ | cAMP: 0 to 3.2 mM: 0, 0.05, 0.1, 0.2, 0.4, 0.8, 1.6, 3.2 mM (dilution by 2-fold) | ||
+ | arabinose: 0 to 0.1%: 0.1, 0.02, 0.004, 0.0008, 0.00016, 0 % (5-fold dilutions) | ||
+ | </section> | ||
+ | |||
</div> | </div> | ||
Revision as of 14:13, 22 August 2012