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Programmable targeted epigenetic editing using CRISPR system in Bombyx mori.

Yue Liu1, Sanyuan Ma2, Jiasong Chang1

  • 1Biological Science Research Center, Southwest University, Chongqing, 400716, China.

Insect Biochemistry and Molecular Biology
|April 26, 2019
PubMed
Summary

Researchers developed a novel tool, dCas9-TET1, to precisely remove DNA methylation in silkworms. This epigenetic modification method enhances gene transcription and aids in understanding insect development.

Keywords:
Bombyx moriCRISPR/dCas9DecitabineDemethylationEpigenetic

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Area of Science:

  • Epigenetics
  • Molecular Biology
  • Genomics

Background:

  • DNA methylation is a key epigenetic mark regulating gene expression in eukaryotes.
  • Its role in insects remains poorly understood due to limited manipulation tools.
  • Efficient tools are needed to investigate DNA methylation's function in insects.

Purpose of the Study:

  • To develop a targeted DNA demethylation tool for the silkworm Bombyx mori.
  • To investigate the functional consequences of locus-specific DNA demethylation in insects.
  • To establish a method for exploring DNA methylation's role in insect development.

Main Methods:

  • Constructed an all-in-one vector expressing a fusion protein of catalytically inactive Cas9 (dCas9) and TET1.
  • Designed guide RNAs (gRNAs) targeting methylated regions in three endogenous silkworm genes.
  • Co-transfected dCas9-TET1 and gRNA into silkworm cells to achieve targeted demethylation.

Main Results:

  • Achieved efficient, target-specific DNA demethylation with efficiencies ranging from 17.50% to 40.00%.
  • Demonstrated that targeted demethylation of gene bodies increased mRNA transcription levels.
  • Showed dCas9-TET1 is more effective and specific than decitabine, with no off-target effects on whole-genome methylation.

Conclusions:

  • dCas9-TET1 provides a powerful and specific tool for locus-specific DNA demethylation in insects.
  • This technology enables functional investigation of DNA methylation in insects.
  • Facilitates exploration of the links between DNA methylation and insect development.