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MicroRNAs01:22

MicroRNAs

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MicroRNA (miRNA) are short, regulatory RNA transcribed from introns (non-coding regions of a gene) or intergenic regions (stretches of DNA present between genes). Several processing steps are required to form biologically active, mature miRNA. The initial transcript, called primary miRNA (pri-mRNA), base-pairs with itself, forming a stem-loop structure. Within the nucleus, an endonuclease enzyme, called Drosha, shortens the stem-loop structure into hairpin-shaped pre-miRNA. After the pre-miRNA...
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MicroRNA (miRNA) are short, regulatory RNA transcribed from introns—non-coding regions of a gene—or intergenic regions—stretches of DNA present between genes. Several processing steps are required to form biologically active, mature miRNA. The initial transcript, called primary miRNA (pri-mRNA), base-pairs with itself forming a stem-loop structure. Within the nucleus, an endonuclease enzyme, called Drosha, shortens the stem-loop structure into hairpin-shaped pre-miRNA. After...
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RNA Interference01:23

RNA Interference

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RNA interference (RNAi) is a process in which a small non-coding RNA molecule blocks the post-transcriptional expression of a gene by binding to its messenger RNA (mRNA) and preventing the protein from being translated.
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Experimental RNAi02:15

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RNA interference (RNAi) is a cellular mechanism that inhibits gene expression by suppressing its transcription or activating the RNA degradation process. The mechanism was discovered by Andrew Fire and Craig Mello in 1998 in plants. Today, it is observed in almost all eukaryotes, including protozoa, flies, nematodes, insects, parasites, and mammals. This precise cellular mechanism of gene silencing has been developed into a technique that provides an efficient way to identify and determine the...
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siRNA - Small Interfering RNAs02:30

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Small interfering RNAs, or siRNAs, are short regulatory RNA molecules that can silence genes post-transcriptionally, as well as the transcriptional level in some cases. siRNAs are important for protecting cells against viral infections and silencing transposable genetic elements.
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Nucleic Acid Structure01:25

Nucleic Acid Structure

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The pentose sugar in DNA is deoxyribose, while in RNA the pentose sugar is ribose. The difference between the sugars is the presence of the hydroxyl group on the ribose's second carbon and a hydrogen on the deoxyribose's second carbon. The phosphate residue attaches to the hydroxyl group of the 5′ carbon of one sugar and the hydroxyl group of the 3′ carbon of the sugar of the next nucleotide, which forms  a 5′ to 3′ phosphodiester linkage.
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Enhanced microRNA accumulation and gene silencing efficiency through optimized precursor base pairing.

Juan-José Llorens-Gámez1, Pedro José García-Cano1, Sara Rico-Rodrigo1

  • 1Instituto de Biología Molecular y Celular de Plantas (Consejo Superior de Investigaciones Científicas-Universitat Politècnica de València), 46022, Valencia, Spain.

The Plant Journal : for Cell and Molecular Biology
|January 8, 2026
PubMed
Summary
This summary is machine-generated.

Optimizing artificial microRNAs (amiRNAs) in plants involves modifying precursor structures. A single G-C pair upstream of the mature amiRNA significantly boosts gene silencing efficiency for crop improvement.

Keywords:
ArabidopsisNicotiana benthamianaRNA silencingartificial microRNAmicroRNA precursor

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Area of Science:

  • Plant molecular biology
  • Gene regulation
  • Biotechnology

Background:

  • MicroRNAs (miRNAs) are key regulators of gene expression in plants, processed from precursors by DICER-LIKE1 (DCL1).
  • Artificial miRNAs (amiRNAs) are tools for targeted gene silencing, but precursor structure optimization is crucial for efficiency.
  • The impact of base pairing near DCL1 cleavage sites on amiRNA production is not well understood.

Purpose of the Study:

  • To investigate how base pairing at or near DCL1 cleavage sites influences amiRNA production from a minimal precursor.
  • To identify structural modifications that enhance amiRNA accumulation and gene silencing efficacy.
  • To validate findings in plant models for potential applications in functional genomics and crop engineering.

Main Methods:

  • Utilized silencing sensor systems in Nicotiana benthamiana to test amiRNA precursor variants.
  • Systematically altered base pairing at naturally mismatched positions near DCL1 cleavage sites.
  • Validated enhanced amiRNA production and silencing in Arabidopsis thaliana transgenic lines using deep sequencing.

Main Results:

  • Introducing a G-C pair upstream of the mature amiRNA significantly enhanced amiRNA accumulation and silencing efficiency.
  • Deep sequencing confirmed accurate processing and predominant release of intended amiRNAs in Arabidopsis.
  • The structural modification demonstrated high specificity and efficacy in enhancing amiRNA-mediated gene silencing.

Conclusions:

  • A single structural modification in amiRNA precursors can substantially improve amiRNA-mediated gene silencing efficacy.
  • The optimized amiRNA platform is suitable for large-scale functional genomics screens.
  • This approach facilitates the development of crops with improved resilience to environmental stresses.