Team:EPF-Lausanne/Our Project/T7 promoter variants/recovery

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(Experimental Setup)
(Results)
 
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We grow two large cultures of cells. One contains cells that will lyse and release plasmids into the supernatant while the other has non-lysing cells containing a control (empty) plasmid. Adding IPTG to both flasks induces lysis in one set of cells but not in the others. If you would like to find out more about how IPTG induction experiments work, please click [[Team:EPF-Lausanne/Our_Project/T7_promoter_variants/lysis/iptg|here]].  
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We grow two large cultures of cells. One contains cells that will lyse and release plasmids into the supernatant while the other has non-lysing cells containing a control (empty) plasmid. Adding IPTG to both flasks induces lysis in one set of cells but not in the others. If you would like to find out more about how IPTG induction experiments work, please click [[EPF-Lausanne/Our_Project/T7_promoter_variants/lysis/iptg|here]].  
[[File:iptg_platecount_text.png|390px|left]]
[[File:iptg_platecount_text.png|390px|left]]
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[[File:firstCFU.png|600px|center]]
[[File:firstCFU.png|600px|center]]
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The colony fluorescence, the second method for quantifying the amount of recovered DNA, relates the number of colonies to the amount of transformant (in uL) that was plated. We find that the number of colonies taken from the lysing culture (all pink because of RFP) increases dramatically over time, while the number of colonies from the non-lysing culture is near-zero over the 10 hours of the experiment. This result is an additional confirmation that DNA was recovered.  
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The colony fluorescence, the second method for quantifying the amount of recovered DNA, relates the number of colonies to the amount of transformant (in uL) that was plated. We find that the number of colonies taken from the lysing culture (all pink because of RFP) increases generally over time, while the number of colonies from the non-lysing culture is near-zero over the 10 hours of the experiment. The colony fluorescence count drops in the last three hours but this phenomenon can be explained by the fact that the contents of the cell (nucleases, etc...) damage the plasmids over time, leading to a lower transformation efficiency. In contrast, the qPCR data shows a monotonic increase in plasmid concentration without a drop over the last few hours. This result is an additional confirmation that DNA was recovered.  
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{{:Team:EPF-Lausanne/Templates/Footer}}

Latest revision as of 18:05, 21 September 2011