Activation of Silent Gene Clusters Mediated by Promoter Substitution

Activation of Silent Gene Clusters Mediated by Promoter Substitution

Promoter substitution is using a new promoter to replace the original promoter of a silenced gene, thus allowing normal expression of the silenced gene in the original host. Since the secondary metabolites of fungi may be toxic to the original host, researchers often use inducible promoters to replace the original promoters of silenced genes to not affect the fungus's growth and activate the silenced genes to obtain new active natural products. However, when performing promoter substitution, the promoters of all silenced biosynthetic genes in the gene cluster need to be replaced, and the workload is high. Lifeasible is committed to developing promoter substitution-mediated activation strategies for silent gene clusters to help customers achieve efficient utilization of fungal resources.

Physical map of the epothilone biosynthetic gene cluster and the promoter Pepo.Figure 1. Physical map of the epothilone biosynthetic gene cluster and the promoter Pepo. (Van Dijck P, et al., 2017)

  • Biosynthetic gene promoter substitution. We use inducible promoters to replace the original promoters of silenced biosynthetic genes and then drive the expression of the target genes in the presence of inducers to synthesize new metabolites.
  • Transcriptional regulatory gene promoter substitution. Fungal biosynthetic gene clusters usually contain multiple transcriptional regulatory genes, which are primarily responsible for synthesizing transcriptional regulators that then specifically bind to the promoter sequences of other genes in the gene cluster, thereby activating the expression of a specific gene cluster.

The specific services we can provide are as follows. Including but not limited to.

Promoter cloning
  • Known promoter sequences
  • Unknown promoter sequences
Promoter activity analysis
  • Report genetic test
Analysis of promoter core elements
  • Promoter 2.0 Prediction Server
  • Softberry
  • Plant CARE
Genetic editing
  • CRISPR/Cas9
Dual fluorescence reporting system
  • Analysis of promoter structure
  • Promoter SNP analysis
  • Interaction analysis of transcription factors and promoters

The commonly used eukaryotic promoters are as follows.

Eukaryotic promoters Types of transcribed RNA Expression features Other features
CMV mRNA Constitutive expression May contain an enhancer that will silence in some cell types
EF1a mRNA Constitutive expression Stable expression in all cell types and physiological states
SV40 mRNA Constitutive expression May contain an enhancer
CAG mRNA Constitutive expression Contains CMV enhancer
UAS mRNA Specific expression Requires the presence of the Gal4 gene to activate the promoter
Ac5 mRNA Constitutive expression Expression systems commonly used in Drosophila
TEF1 mRNA Constitutive expression Similar to the mammalian EF1α promoter
GDS mRNA Constitutive expression Great ability to start expressions
ADH1 mRNA Inhibition with ethanol This promoter has a strong initiation ability and high expression level when it is full-length; it is constitutively expressed when it is truncated and has a low expression level

Lifeasible can provide full operational services for promoter substitution-mediated activation of silent gene clusters, helping our customers to reduce project time. As your trusted partner, we can meet all your fungal phylogenetic analysis needs and provide efficient, high-quality services. If you want to know the details, please contact us.

Reference

  1. Yue XJ, et al. Effects of transcriptional mode on promoter substitution and tandem engineering for the production of epothilones in Myxococcus xanthus. Appl Microbiol Biotechnol. 2018 Jul; 102(13): 5599-5610.
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