Part:BBa_K2259074

Rop protein with anderson (0.24) promoter for global plasmid copy number control

This construct is an intermediate to full SynORI global copy number control device. It consists of a weak Anderson promoter, weak Anderson RBS, ROP protein and a double terminator.

When placed in a SynORI system this device lowers plasmid copy number of each group as it can bypass the selective control.

Device also works with other plasmids consisting of ColE1 origin of replication

Devices from the same series that have different Anderson promoters: part:BBa_K2259072 (0 Anderson), part:BBa_K2259073 (0.15 Anderson), part:K2259074 (0.24 Anderson).

See how this part fits into the whole SynORI framework by pressing here!

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Figure 1. Main principles of ColE1 plasmid family replication. Rop protein interaction region marked in red square. (Citation needed)

Introduction

Biology

Repressor of primer (ROP) is a small dimeric protein that participates in ColE1 plasmid family copy number control, by increasing affinity between two complementary RNAs - RNA I (Replication inhibitor) and RNA II (Replication activator) (Fig. 1). By increasing affinity of the two RNA molecules, Rop decreases the rate of plasmid replication initiation events.

Figure 2. Structure of the ColE1 Rop protein, at 1.7 angstroms resolution.

Rop dimer is a bundle of four tightly packed alpha helices that are held by hydrophobic interactions (Fig. 2).

Usage with SynORI (Framework for multi-plasmid systems)

About SynORI

SynORI is a framework for multi-plasmid systems created by Vilnius-Lithuania 2017 which enables quick and easy workflow with multiple plasmids, while also allowing to freely pick and modulate copy number for every unique plasmid group! Read more about [http://2017.igem.org/Team:Vilnius-Lithuania SynORI here]!

This device in SynORI

This is a constitutive global copy number modulator device which lowers plasmid copy number of every group in the system bypassing the selective control of different groups. These constitutive devices can be used with different Anderson promoters to select a different copy number.

Devices from the same series that have different Anderson promoters: part:BBa_K2259072 (0 Anderson), part:BBa_K2259073 (0.15 Anderson), part:K2259074 (0.24 Anderson).

See the [http://2017.igem.org/Team:Vilnius-Lithuania Vilnius-Lithuania 2017 team wiki] for more insight about our synthetic origin of replication (SynORI).

Further details

For more background information and indepth insight on this part's design please see the individual part page of ROP proteinpart:BBa_K2259010.

Characterization

In order to characterize this construct, it must be cloned next to RNA II gene. Please see part:BBa_K2259052.