Difference between revisions of "Part:BBa K1741000"

Revision as of 20:26, 14 October 2016

sfGFP under arabinose promoter s1 without AraC (Arashort1)

In commercially available expression vectors arabinose promoter araBAD is usually used together with a gene for AraC transcription activator/repressor to make the system independent of chromosomal copy of AraC gene. We fused the araBAD promoter without AraC reading frame to sfGFP as well as other sugar induced promoters. The shorter version of the popular promoter is still functional i.e. induced by arabinose. Arabinose (0,4%) induced expression is a bit lower than driven by AraC-araBAD after a short time (1-8h) but after a longer time like 18h the GFP level is higher than from AraC-araBAD promoter. New shorter versions of arabinose induced promoters, all originating from E. coli genome were compared to the biobrick BBa_K1481002, provided last year by Poznan_Bioinf team.

Contribution

Group: UAM_Poznan 2016 Authors: Przemysław Nuc Summary:

Design

Legend

AraWT - [BBa_K1481002]

Arashort1 - [BBa_1741000]

Arashort2 - [BBa_1741002]

For the design of our modified arabinose promoters we used the BioBrick created by our fellow iGEM team - PoznanBioInf. Their promoter was used as a control for out other arabinose promoters. We shortened the sequence of the original promoter by removing lengthy AraC part (we called this construct Arashort1) or removing AraC together with operator regions O1 and O2 (we called this construct Arashort2).

Characteristics and results

Legend

AraWT - [BBa_K1481002]

Arashort1 - [BBa_1471000]

Arashort2 - [BBa_1471002]

The sfGFP fluorescence [RFU] was measured using Tecan fluorometer.

All of our arabinose promoters are induced by the arabinose and repressed by the glucose. Arashort1 works better than AraWT. On the other hand, Arashort2 is slightly weaker when compared to Arashort1, but still stronger than AraWT. Probably O1 and O2 regions that were removed in Arashort2 are necessary for the best performance of the promoter.

We also checked the tightness of our promoters.

All of our promoters are induced only by their respective sugars with a small exception of xylose promoters being slightly induced by arabinose and arabinose promoters being slightly induced by xylose. All of our promoters are repressed by glucose.

Sequence and Features