Difference between revisions of "Part:BBa K1618037"

Latest revision as of 16:26, 21 September 2015

35s Terminator

35S is a plant specific 3' untranslated sequence and terminator from Cauliflower Mosaic Virus. The part’s intended use is to terminate transcription in plant cells. This part was converted to a standard BioBrick by cloning into the Mo-Flipper Plasmid K1467300 made by the NRP-UEA 2014 team.

NRP-UEA 2015 team used this part in our plant constructs. Constructs K1618029-36 contained a 35s promoter, chloroplast transit peptide, one of four acyltransferases, and this new 35s terminator parts. In parts K1618029-32 fused the recombinant protein was fused to a fluorescent a reporter tag to confirm the localisation to the chloroplast (Figure 1). The parts were tested by transferring into Agrobacterium tumefaciens and infiltration into plants. The correct and strong expression indicates that the terminator is functional.

Figure 1: Constructs BBa_K1618029‐032 contain a yellow fluorescent protein, as well as a chloroplast transit peptide. These are confocal microscopy images of the constructs infiltrated into Nicotiana benthamiana, in which the red structures are the chlorophyll within the chloroplast, and the yellow is the fluorescent fusion protein expressed from constructs (a) BBa_K1618029, (b) BBa_K1618031, (c) BBa_K1618032, and (d) BBa_K1618030.

The above results not only indicate a successful localisation, but also suggest that the 35s terminator that we used is GoldenGate compatible and functioning.

Secondary Structure

File:Mfold-K1618037-1.png

Measurement

[http://openwetware.org/wiki/Cconboy:Terminator_Characterization/Results How these parts were measured]

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-S is a plant specific terminator obtained from the Cauliflower Mosaic Virus. The part’s intended use is to terminate transcription in plant cells. This part was converted to a standard BioBrick by cloning into the Mo-Flipper Plasmid K1467300 made by the NRP-UEA 2014 team.
+S is a plant specific 3' untranslated sequence and terminator from  Cauliflower Mosaic Virus. The part’s intended use is to terminate transcription in plant cells. This part was converted to a standard BioBrick by cloning into the Mo-Flipper Plasmid K1467300 made by the NRP-UEA 2014 team.
-NRP-UEA 2015 team used this part in our plant constructs. Constructs K1618033-36 contained a 35s promoter, chloroplast transit peptide, one of four acyltransferase, and this 35s terminator. Parts K1618029-32 were the same as the first four, but also contained a GFP tag to confirm the localisation to the chloroplast.
-We transformed our constructs into <i>Agrobacterium tumefaciens</i> and then infiltrated it into our plants. We used the YFP tagged constructs to confirm the localisation of parts to the chloroplast (Figure 1) and the untagged constructs to analyse the starch content of our leaves.
+NRP-UEA 2015 team used this part in our plant constructs. Constructs K1618029-36 contained a 35s promoter, chloroplast transit peptide, one of four acyltransferases, and this new 35s terminator parts. In parts K1618029-32 fused the recombinant protein was fused to a fluorescent a reporter tag to confirm the localisation to the chloroplast (Figure 1). The parts were tested by transferring into <i>Agrobacterium tumefaciens</i> and infiltration into plants. The correct and strong expression indicates that the terminator is functional.