Difference between revisions of "Part:BBa K3171172"

Revision as of 10:45, 21 October 2019

Vibrio natriegens Native P1 promotor with mCherry

Vibrio natriegens has been reported to have a remarkable doubling time of 9.8 min. As is the case for the better-studied but slower-growing E. coli, V. natriegens increases its number of ribosomes with the growth rate in order to achieve its extraordinarily high rate of protein synthesis. Multiple mechanisms contribute to this high ribosome synthesis efficiency, including high rRNA gene copy number, strong promoters that contain near-consensus −10, −35, and UP elements and activation by the transcription factor Fis. In addition, V. natriegens rRNA promoters exhibit the relatively short-lived open-complex characteristic of rRNA promoters in E. coli, potentially contributing to the regulation of these promoters in vivo. The native promoter P1 is a constitutive promoter. We fused the the native P1 promoter with mCherry fluorescent protein with the help of the 3A assembly cloning method.

The P1 promoter with Biobrick prefix and suffix was cloned with mCherry to obtain a composite part that constitutively expresses mCherry Part:BBa_J06504. The P1-mCherry plasmid was cloned into a backbone plasmid of Chloramphenicol resistance (pSB1C3).

Genes under the control of this promoter are constitutively transcribed by the organism’s ribosomal protein P1. We show that there is a 225 times increase of fluorescence using the P1 promoter in E. coli compared to E. coli without a plasmid. At the same time, measurement in V. natriegens showed a 3 times increase of fluorescence (figure 1).

Figure 1: Fluorescence of the reporter mCherry under control of the constitutive P1 promoter in V. natriegens and E. coli in comparison to control without plasmid.

Usage and Biology

Sequence and Features