BBa_K731700 1 BBa_K731700 Platform for terminators analysis under the control of T7 promoter 2012-08-13T11:00:00Z 2015-05-08T01:13:06Z It's made from a Roberta Lentini's construct (from Mansy lab) that consist of pET21b with the two proteins and a 20bp linker between them. That construct was mutated two times to eliminate illegal restriction sites; prefix-suffix linker was added by insertion PCR. We used this part to test terminators: the two subsequent proteins allow to quantify the termination efficiency as the ratio between Venus's and Cherry's fluorescence. The vector is pET21b, that contains lacI, t7 promoter and ampicillin resistance gene. The two fluorescent proteins have partially overlapping emission spectra, thus the fluorimetric measures should be taken with these parameters, to avoid interference between the two: Venus excitation: 485nm, Venus emission: 528nm Cherry excitation: 528nm, Cherry emission: 615nm We inserted the prefix-suffix linker between the two proteins, to allow terminator characterization. false false _977_ 0 12063 9 It's complicated true Using and modifying this construct is important to remember that the two proteins have strong N-terminal homology, especially designing primers. For any future improvement, maybe it would be better two use two different fluorescence proteins, because these two has partially overlapping emission spectra and different folding kinetics. Using and modifying this construct is important to remember that the two proteins have strong N-terminal homology, especially designing primers. For any future improvement, maybe it would be better two use two different fluorescence proteins, because these two has partially overlapping emission spectra and different folding kinetics. false Giacomo Giacomelli, Anna Depetris annotation2179559 1 lacI range2179559 1 4562 5653 annotation2179560 1 T7 PROMOTER range2179560 1 6031 6050 annotation2179556 1 A206K VENUS range2179556 1 36 752 annotation2179557 1 T7 TERMINATOR range2179557 1 843 890 annotation2179558 1 ampicillin resistance gene range2179558 1 1711 2370 annotation2179562 1 mCHERRY range2179562 1 6119 6829 annotation2179561 1 lacO range2179561 1 6050 6072 BBa_K731700_sequence 1 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igem2sbol 1 iGEM to SBOL conversion Conversion of the iGEM parts registry to SBOL2.1 James Alastair McLaughlin Chris J. Myers 2017-03-06T15:00:00.000Z