Simple and rapid direct cloning and heterologous expression of natural product biosynthetic gene cluster in Bacillus subtilis via Red/ET recombineering

Liu, Qingshu; Shen, Quanli; Bian, Xiaoying; Hanna, Calvin; Fu, Jun; Wang, Hailong; Lei, Ping; Guo, Zhaohui; Wu, Canrong; Li, Dingjun; Zhang, Youming

doi:10.1038/srep34623

Cited by 34 publications

(39 citation statements)

References 48 publications

(86 reference statements)

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“…As with the actinomycetes, Red/ET‐based recombineering methods have also been used for the heterologous expression of the bacillomycin BGC from B. amyloliquefaciens (37.2kb) in B. subtilis , showing a rapid (<1 week) method to express large gene clusters and improving the workflow of heterologous expression in this organism. However, this approach only showed success with one of the two investigated BGCs in the paper (the other being the cluster for edeine production), and so may not be suitable for all classes of Bacillus ‐derived antimicrobials …”

Section: Chassismentioning

confidence: 97%

“…Approaches used for the expression of antibiotic BGCs in non‐Streptomyces organisms. Methods include combinatorial DNA assembly of benzylpenicillin in Saccharomyces cerevisiae , Red/ET recombineering, and TAR cloning in Bacillus subtilis . In the shown example of combinatorial assembly, a library of pathway genes attached to promoters of varying strength is generated.…”

Section: Chassismentioning

confidence: 99%

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The “Three Cs” of Novel Antibiotic Discovery and Production through Synthetic Biology: Biosynthetic Gene Clusters, Heterologous Chassis, and Synthetic Microbial Consortia

2018

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Antimicrobial resistance is a rapidly growing problem worldwide, with bacteria showing resistance to last‐resort antibiotic treatments such as carbapenems and colistin becoming more abundant. Synthetic biology may be able to contribute to solving this growing antimicrobial resistance crisis by facilitating the engineering of diversified collections of novel antibiotics, targeting multidrug resistant bacteria in new ways. This review discusses recent advances in the use of synthetic biology methodologies to discover and produce novel antimicrobials; in particular, the work being carried out on biosynthetic gene clusters, heterologous chassis, and synthetic microbial consortia, the “three Cs” of the title.

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Section: Chassismentioning

confidence: 97%

Section: Chassismentioning

confidence: 99%

The “Three Cs” of Novel Antibiotic Discovery and Production through Synthetic Biology: Biosynthetic Gene Clusters, Heterologous Chassis, and Synthetic Microbial Consortia

2018

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“…Luminmide A/B (Fu et al, 2012) Bacillomycin (Liu Q. et al, 2016) Streptoketides (Qian et al, 2020) ExoCET…”

Section: Llhrunclassified

Bioprospecting Through Cloning of Whole Natural Product Biosynthetic Gene Clusters

Lin

Nielsen

Liu

2020

Front. Bioeng. Biotechnol.

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Since the discovery of penicillin, natural products and their derivatives have been a valuable resource for drug discovery. With recent development of genome mining approaches in the post-genome era, a great number of natural product biosynthetic gene clusters (BGCs) have been identified and these can potentially be exploited for the discovery of novel natural products that can find application as pharmaceuticals. Since many BGCs are silent or do not express in native hosts under laboratory conditions, heterologous expression of BGCs in genetically tractable hosts becomes an attractive route to activate these BGCs to discover the corresponding products. Here, we highlight recent achievements in cloning and discovery of natural product biosynthetic pathways via intact BGC capturing, and discuss the prospects of high-throughput and multiplexed cloning of rational-designed gene clusters in the future.

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“…Bandage Tool for visualizing de Bruijn assembly graphs, allows for a deeper analysis of de novo assemblies which is not accessible through study of individual contigs. [50] pathways such as the NPRS clusters coding for edeine (48.3 kb) and bacillomycin (37.3 kb) [70].…”

Section: Grapementioning

confidence: 99%

Exploration and exploitation of the environment for novel specialized metabolites

Loureiro

Medema

Oost

et al. 2018

Current Opinion in Biotechnology

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Microorganisms are Nature's little engineers of a remarkable array of bioactive small molecules that represent most of our new drugs. The wealth of genomic and metagenomic sequence data generated in the last decade has shown that the majority of novel biosynthetic gene clusters (BGCs) is identified from cultivation-independent studies, which has led to a strong expansion of the number of microbial taxa known to harbour BGCs. The large size and repeat sequences of BGCs remain a bioinformatic challenge, but newly developed software tools have been created to overcome these issues and are paramount to identify and select the most promising BGCs for further research and exploitation. Although heterologous expression of BGCs has been the greatest challenge until now, a growing number of polyketide synthase (PKS) and non-ribosomal peptide synthetase (NRPS)-encoding gene clusters have been cloned and expressed in bacteria and fungi based on techniques that mostly rely on homologous recombination. Finally, combining ecological insights with state-of-the-art computation and molecular methodologies will allow for further comprehension and exploitation of microbial specialized metabolites.

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Simple and rapid direct cloning and heterologous expression of natural product biosynthetic gene cluster in Bacillus subtilis via Red/ET recombineering

Cited by 34 publications

References 48 publications

The “Three Cs” of Novel Antibiotic Discovery and Production through Synthetic Biology: Biosynthetic Gene Clusters, Heterologous Chassis, and Synthetic Microbial Consortia

The “Three Cs” of Novel Antibiotic Discovery and Production through Synthetic Biology: Biosynthetic Gene Clusters, Heterologous Chassis, and Synthetic Microbial Consortia

Bioprospecting Through Cloning of Whole Natural Product Biosynthetic Gene Clusters

Exploration and exploitation of the environment for novel specialized metabolites

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