Difference between revisions of "Team:UNSW Australia/Demonstrate"

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                 <h2 style="font-family: Slim-Joe;">Demonstration of Hypervesiculation</h2>
 
                 <h2 style="font-family: Slim-Joe;">Demonstration of Hypervesiculation</h2>
 
                 <p>Our results showed that the knock-in mutation of g3p successfully induced hypervesiculation of E. coli with an OMV count of 2.8x108 per mL per OD600, significantly higher than the count obtained from the negative control sample (0.29x108 per mL per OD600) (Figure 1). Knock-out of TolA also showed an impressive increase in OMV production, over 36 times the amount in wildtype E. coli (Figure 1). This affect was all the more pronounced when TolR overexpression was added to this same knockout strain (Figure 1). Knockout of DegP also resulted in increased OMV production compared to the wildtype, nevertheless production fell very short of the production rates seen in the other mutants. Most importantly, however, knockout of DegP was associated with a dramatic increase in vesicle size, if not vesicle number (Figure 2).</p>
 
                 <p>Our results showed that the knock-in mutation of g3p successfully induced hypervesiculation of E. coli with an OMV count of 2.8x108 per mL per OD600, significantly higher than the count obtained from the negative control sample (0.29x108 per mL per OD600) (Figure 1). Knock-out of TolA also showed an impressive increase in OMV production, over 36 times the amount in wildtype E. coli (Figure 1). This affect was all the more pronounced when TolR overexpression was added to this same knockout strain (Figure 1). Knockout of DegP also resulted in increased OMV production compared to the wildtype, nevertheless production fell very short of the production rates seen in the other mutants. Most importantly, however, knockout of DegP was associated with a dramatic increase in vesicle size, if not vesicle number (Figure 2).</p>
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                <p>Figure 1</p>
 
 
                 <img class="img-responsive" src="https://static.igem.org/mediawiki/parts/3/34/T--UNSW_Australia--ParticleCount.png">
 
                 <img class="img-responsive" src="https://static.igem.org/mediawiki/parts/3/34/T--UNSW_Australia--ParticleCount.png">
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                 <p>Figure 2</p>
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                 <p><i><b>Figure 1<b> OMV count x 10^8, per mL, per OD600, as measured by NanoSight LM20. Samples are: Control = NEB T7 E. coli; g3p (non-induced) = NEB T7 + g3p; g3p (induced) = NEB T7 E. coli + g3p + 1mM IPTG; ΔdegP = ΔdegP E. coli; ΔtolA = ΔtolA E. coli; ΔtolA + tolR (induced) = ΔtolA E. coli + TolR + 50mM arabinose</i></p>
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                 <img class="img-responsive" src="https://static.igem.org/mediawiki/2016/1/14/T--UNSW_Australia--degp_results.png">
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                 <img class="img-responsive" src="https://static.igem.org/mediawiki/2016/1/14/T--UNSW_Australia--degp_results.png">
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                <p><i><b>Figure 2</b>Demonstrate a wide distribution of vesicle sizes produced<i></p>
 
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             </div>
 
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Revision as of 03:27, 20 October 2016

Demonstration of Hypervesiculation

Our results showed that the knock-in mutation of g3p successfully induced hypervesiculation of E. coli with an OMV count of 2.8x108 per mL per OD600, significantly higher than the count obtained from the negative control sample (0.29x108 per mL per OD600) (Figure 1). Knock-out of TolA also showed an impressive increase in OMV production, over 36 times the amount in wildtype E. coli (Figure 1). This affect was all the more pronounced when TolR overexpression was added to this same knockout strain (Figure 1). Knockout of DegP also resulted in increased OMV production compared to the wildtype, nevertheless production fell very short of the production rates seen in the other mutants. Most importantly, however, knockout of DegP was associated with a dramatic increase in vesicle size, if not vesicle number (Figure 2).

Figure 1 OMV count x 10^8, per mL, per OD600, as measured by NanoSight LM20. Samples are: Control = NEB T7 E. coli; g3p (non-induced) = NEB T7 + g3p; g3p (induced) = NEB T7 E. coli + g3p + 1mM IPTG; ΔdegP = ΔdegP E. coli; ΔtolA = ΔtolA E. coli; ΔtolA + tolR (induced) = ΔtolA E. coli + TolR + 50mM arabinose

Figure 2Demonstrate a wide distribution of vesicle sizes produced