Team:Bologna/Project

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Introduction

Our goal is to create a logic gate based on a post-transcriptional regulation system in Escherichia coli, using RNA to silence gene expression. We inserted a cis-acting sequence directly upstream of the rbs (ribosome binding site) to realize a "regulated rbs". We also designed the complementary trans-repressing sequence whose function is to recognize and cover the "regulated rbs" and prevent translation from it. Two versions of the trans- repressing sequence were designed with 2 different kinds of rbs covers. We want to use this short non-coding RNA segment placed in trans, to silence translation from dowstream the cis-acting sequence. We developed a bioinformatic tool to research the sequences and these are the results:

Cis-repressing sequence, inserted upstream of the target gene
Scar Cis-repressing Rbs Scar
TACTAGAG AACACAAACTATCACTTTAACAACACATTACATATACATTAAAATATTAC AAAGAGGAGAAA TACTAGAG


Trans-repressor sequence with a cover of 7 bases (long version)
Scar Cover Trans(long) Scar
TACTAGAG CCTCTTT GTAATATTTTAATGTATATGTAATGTGTTGTTAAAGTGATAGTTTGTGTT TACTAGAG


Trans-repressor sequence with a cover of 4 bases (short version)
Scar Cover Trans(short) Scar
TACTAGAG CTTT GTAATATTTTAATGTATATGTAATGTGTTGTTAAAGTGATAGTTTGTGTT TACTAGAG



The sequence secondary structures of RNAs are obtained using the RNA mfold tool:

Figure 1: Cis-repressing sequence
Figure 2: Trans-repressor sequence (long version)
Figure 3: Trans-repressor sequence (short version)


These are the three genetic circuits that we are currently studying:

Circuito with rRBS 1.jpg


Circuito with rRBS 1.1.jpg



Circuito with rRBS 2.jpg