Novel methods to optimize gene and statistic test for evaluation – an application for Escherichia coli

Tran, Tuan Anh; Ly, Le Thi Thuy; Viet, Ngo Quoc; Bao, Pham The

doi:10.1186/s12859-017-1517-z

Cited by 4 publications

(7 citation statements)

References 43 publications

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“…To ensure efficient translation of synthetic genes in the host organism Mutation Maker also features reverse translation functionality that is tightly coupled to codon optimization. In this regard several proprietary and freely available codon optimization algorithms have been developed. ,,− However, many of the published methods optimize the codons in the target gene with respect to the CAI . The CAI reflects the deviation of codon frequencies in a given sequence from those observed in a reference set of highly expressed genes in the target host organism.…”

Section: Discussionmentioning

confidence: 99%

Mutation Maker, An Open Source Oligo Design Platform for Protein Engineering

Hiraga

Mejzlík²,

Marcisin³

et al. 2021

ACS Synth. Biol.

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Protein engineering is the discipline of developing useful proteins for applications in research, therapeutic, and industrial processes by modification of naturally occurring proteins or by invention of de novo proteins. Modern protein engineering relies on the ability to rapidly generate and screen diverse libraries of mutant proteins. However, design of mutant libraries is typically hampered by scale and complexity, necessitating development of advanced automation and optimization tools that can improve efficiency and accuracy. At present, automated library design tools are functionally limited or not freely available. To address these issues, we developed Mutation Maker, an open source mutagenic oligo design software for large-scale protein engineering experiments. Mutation Maker is not only specifically tailored to multisite random and directed mutagenesis protocols, but also pioneers bespoke mutagenic oligo design for de novo gene synthesis workflows. Enabled by a novel bundle of orchestrated heuristics, optimization, constraint-satisfaction and backtracking algorithms, Mutation Maker offers a versatile toolbox for gene diversification design at industrial scale. Supported by in silico simulations and compelling experimental validation data, Mutation Maker oligos produce diverse gene libraries at high success rates irrespective of genes or vectors used. Finally, Mutation Maker was created as an extensible platform on the notion that directed evolution techniques will continue to evolve and revolutionize current and future-oriented applications.

show abstract

Section: Discussionmentioning

confidence: 99%

Mutation Maker, An Open Source Oligo Design Platform for Protein Engineering

Hiraga

Mejzlík²,

Marcisin³

et al. 2021

ACS Synth. Biol.

View full text Add to dashboard Cite

show abstract

“…To ensure efficient translation of synthetic genes in the host organism Mutation Maker also features reverse translation functionality that is tightly coupled to codon optimization. In this regard several proprietary and freely available codon optimization algorithms have been developed (Chung and Lee, 2012; Gould et al, 2014; Liu et al, 2014; Raab et al, 2010; Swainston et al, 2014; Tuan-Anh et al, 2017). However, many of the published methods optimize the codons in the target gene with respect to the CAI (Sharp and Li, 1987).…”

Section: Discussionmentioning

confidence: 99%

Mutation Maker, An Open Source Oligo Design Platform for Protein Engineering

Hiraga

Mejzlík²,

Marcisin³

et al. 2020

Preprint

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Founded on principles of natural selection, directed evolution techniques harness high-throughput gene diversification methodologies to generate proteins with optimized properties for applications in research, therapeutic and industrial processes. However, design of gene libraries is typically hampered by scale and complexity, necessitating development of advanced automation and optimization tools that can improve efficiency and accuracy. At present, automated library design tools are obsolete, functionally limited or not freely available for commercial use. To address these issues, we developed Mutation Maker, an open source mutagenic oligo design software for large-scale protein engineering experiments. Mutation Maker is not only specifically tailored to multi-site random and directed mutagenesis protocols, but also pioneers bespoke mutagenic oligo design for de novo gene synthesis workflows. Enabled by a novel bundle of orchestrated heuristics, optimization, constraint-satisfaction and backtracking algorithms, Mutation Maker offers a versatile toolbox for gene diversification design at industrial scale. Supported by in-silico simulations and compelling experimental validation data, Mutation Maker oligos produce diverse gene libraries at high success rates irrespective of genes or vectors used. Finally, Mutation Maker was created as an extensible platform on the notion that directed evolution techniques will continue to evolve and revolutionize current and future-oriented applications.

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“…Cap-SpyCatcher (Cap-SC) construct includes an N-terminal 6×His tag, PCV2 Cap (amino acids 16 to 233), a flexible linker (GGGGS), and a truncated SpyCatcher [ 34 ] at C-terminal. After optimization of E.coli codon bias for enhanced protein expression [ 35 , 36 ], it was synthesized by Hangzhou Tsingke Biotechnology Company. DNA sequence encoding SpyTag (AHIVMVDAYKPTK) was fused to N-terminus of sfGFP (GenBank, ASL68970.1) by PCR to form ST-sfGFP construct (named ST-GFP).…”

Section: Methodsmentioning

confidence: 99%

A modular and self-adjuvanted multivalent vaccine platform based on porcine circovirus virus-like nanoparticles

et al. 2022

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Background Virus-like particles (VLPs) are supramolecular structures composed of multiple protein subunits and resemble natural virus particles in structure and size, making them highly immunogenic materials for the development of next-generation subunit vaccines. The orderly and repetitive display of antigenic epitopes on particle surface allows efficient recognition and cross-link by B cell receptors (BCRs), thereby inducing higher levels of neutralizing antibodies and cellular immune responses than regular subunit vaccines. Here, we present a novel multiple antigen delivery system using SpyCatcher/Spytag strategy and self-assembled VLPs formed by porcine circovirus type 2 (PCV2) Cap, a widely used swine vaccine in solo. Results Cap-SC, recombinant Cap with a truncated SpyCatcher polypeptide at its C-terminal, self-assembled into 26-nm VLPs. Based on isopeptide bonds formed between SpyCatcher and SpyTag, classical swine fever virus (CSFV) E2, the antigen of interest, was linked to SpyTag and readily surface-displayed on SpyCatcher decorated Cap-SC via in vitro covalent conjugation. E2-conjugated Cap VLPs (Cap-E2 NPs) could be preferentially captured by antigen presenting cells (APCs) and effectively stimulate APC maturation and cytokine production. In vivo studies confirmed that Cap-E2 NPs elicited an enhanced E2 specific IgG response, which was significantly higher than soluble E2, or the admixture of Cap VLPs and E2. Moreover, E2 displayed on the surface did not mask the immunodominant epitopes of Cap-SC VLPs, and Cap-E2 NPs induced Cap-specific antibody levels and neutralizing antibody levels comparable to native Cap VLPs. Conclusion These results demonstrate that this modularly assembled Cap-E2 NPs retains the immune potential of Cap VLP backbone, while the surface-displayed antigen significantly elevated E2-induced immune potency. This immune strategy provides distinctly improved efficacy than conventional vaccine combination. It can be further applied to the development of dual or multiple nanoparticle vaccines to prevent co-infection of PCV2 and other swine pathogens.

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Novel methods to optimize gene and statistic test for evaluation – an application for Escherichia coli

Cited by 4 publications

References 43 publications

Mutation Maker, An Open Source Oligo Design Platform for Protein Engineering

Mutation Maker, An Open Source Oligo Design Platform for Protein Engineering

Mutation Maker, An Open Source Oligo Design Platform for Protein Engineering

A modular and self-adjuvanted multivalent vaccine platform based on porcine circovirus virus-like nanoparticles

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