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Review
. 2021 Sep 15;6(4):254-261.
doi: 10.1016/j.synbio.2021.09.003. eCollection 2021 Dec.

Synthetic polycistronic sequences in eukaryotes

Xuekun Wang  1 , Mario Andrea Marchisio  1
Affiliations
Review

Synthetic polycistronic sequences in eukaryotes

Xuekun Wang et al. Synth Syst Biotechnol. .

Abstract

The need for co-ordinate, high-level, and stable expression of multiple genes is essential for the engineering of biosynthetic circuits and metabolic pathways. This work outlines the functionality and design of IRES- and 2 A-peptide-based constructs by comparing different strategies for co-expression in polycistronic vectors. In particular, 2 A sequences are small peptides, mostly derived from viral polyproteins, that mediate a ribosome-skipping event such that several, different, separate proteins can be generated from a single open reading frame. When applied to metabolic engineering and synthetic gene circuits, 2 A peptides permit to achieve co-regulated and reliable expression of various genes in eukaryotic cells.

Keywords: 2A peptide; IRES; Polycistronic sequence; Synthetic biology.

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Conflict of interest statement

The authors Xuekun Wang and Mario Andrea Marchisio declare no conflict of interest.

Figures

Fig. 1
Fig. 1
The translation mechanism of IRES-based bicistronic constructs.
Fig. 2
Fig. 2
The translation mechanism of 2 A-based bicistronic constructs.
Fig. 3
Fig. 3
Bi-cistronic constructs with different 2 A peptides alone and combined either in tandem or in triplets.
Fig. 4
Fig. 4
The AND gate based on dCas9-mediated transcription activation. There are two different reporter systems: single reporter system using a yellow fluorophore (Venus) as reporter and double reporter system containing a combination of GOI (gene of interest) and GFP via T2A peptide. The pGAL and LexA-ER (estrogen receptor)-AD (activation domain) were activated by inducers such as galactose and β-estradiol, respectively, and led to the expression of MCP-VP64 and dCas9. Only in the presence of both input molecules reporter (and GOI) gene expression takes place
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