Team:Evry/Interlab Study/Used Devices

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IGEM Evry 2014

Interlab study - Used devices

Used Biobricks and plasmids

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Promoter name, plate and well location into the iGEM distribution kit 2014, vector, resistance cassette, part size in bp, used primers and PCR product profile.

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Plasmid map 1: PSB1C3 map with restriction sites (NotI, BsaI, EcoRI, PstI, SpeI and XbaI)

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Plasmid map 2: BBa_J61002 plasmid map with restriction sites ( EcoRI, PstI, SpeI and XbaI)

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Plasmid map 3: BBa_J61002 plasmid map with restriction sites and J23101 promoter ( EcoRI, PstI, SpeI and XbaI)

Construction way

To test the expression of promoters, they are cloned into the PSB1C3 vector followed by the BBa_E0240, a GFP generator. Constructions are inserted in E. coli DH5 alpha that grown on LB medium added with the selection agent, chloramphenicol.

Analysis protocol

Our team decided to test the fluorescence with a TECAN infiniteM200 in 96 well plates.
Fluorescence data were analyzed via following formula that come from 2010 De Jong et al Experimental and computational validation of models of fluorescent and luminescent reporter genes in bacteria .

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Equation A: A(t) is the corrected absorbance of DH5 alpha with the functional reporter system, Au(t)the uncorrected absorbance of DH5 alpha with the functional reporter system and Ab(t)the background absorbance corresponding to LB with chloramphenicol medium absorbance.
Equation B: B(t) is the absorbance of the strain carrying the promoterless vector, Iu(t) the uncorrected fluorescence intensity of DH5 alpha with the functional reporter system and Ib(t) the background fluorescence intensity of LB chloramphenicol medium with the DH5 alpha carrying the promoterless vector.

Contacts

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