View source for Part:BBa K3032127


__NOTOC__
<partinfo>BBa_K3032127 short</partinfo>

The system based on re-engineered ColE1 origin of replication allowing to regulate plasmid copy number in E. coli cells with light. ColE1 plasmid replicon is based on two antisense RNA molecules: RNA I and RNA II. RNA II transcript of forms a RNA-DNA duplex with plasmid and acts as a primer for DNA polymerase. For that reason, RNA II is often called a replication initiator. However, another molecule - RNA I may bind to its antisense version of RNA II, which results in replication inhibition.

Jayraman et al. (2016) engineered a novel bidirectional promoter system for Escherichia coli that can be induced or repressed rapidly and reversibly using the blue light dependent DNA-binding protein EL222. Inserting EL222 binding sequence between -35 and -10 promoter regions led to greater than 3-fold reduction in RFP fluorescence when the engineered E. coli was exposed to blue light.

Vilnius-Lithuania iGEM 2019 combined RNA I/RNA II with EL222 protein to develop light-regulated system for plasmid copy number control. EL222 binding sequence was inserted between -35 and -10 regions of strong Anderson promoter (BBa_J23102), controlling transcription of the replication inhibiting – RNA I molecule. In blue light illumination EL222 protein binds to RNA I promoter. Inhibited transcription results reduced formation of RNA II-RNA I duplex and expanded plasmid copy number.

<!-- Add more about the biology of this part here
===Usage and Biology===

<!-- -->
<span class='h3bb'>Sequence and Features</span>
<partinfo>BBa_K3032127 SequenceAndFeatures</partinfo>


<!-- Uncomment this to enable Functional Parameter display 
===Functional Parameters===
<partinfo>BBa_K3032127 parameters</partinfo>
<!-- -->