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Related Concept Videos

MicroRNAs01:22

MicroRNAs

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 ends...
MicroRNAs01:22

MicroRNAs

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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microRNAs for qPCR Normalization Under Morphofunctional Conditions in Bovine Sperm (Bos taurus).

Lucas Petitemberte de Souza1, Leandro Silva Nunes1, Luana Carla Salvi1

  • 1Laboratório de Genômica Estrutural, Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, Brasil.

Molecular Reproduction and Development
|August 6, 2025
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Summary

Identifying the best microRNAs (miRNAs) for normalizing quantitative polymerase chain reaction (qPCR) in cattle semen is crucial for accurate fertility assessments. This study found specific miRNAs suitable for analyzing sperm motility and morphology, improving cattle breeding efficiency.

Keywords:
Bos taurusnormalizing microRNAsqPCRsemenstability

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Area of Science:

  • Reproductive biology and livestock science.
  • Molecular biology and gene expression analysis.
  • Quantitative genetics and bioinformatics.

Background:

  • Cattle fertility is vital for successful livestock production.
  • Sperm microRNAs (miRNAs) regulate sperm function, but their accurate quantification requires reliable normalization.
  • Current methods lack universally accepted reference miRNAs for bovine semen analysis.

Purpose of the Study:

  • To identify and validate stable microRNA (miRNA) normalizers for quantitative polymerase chain reaction (qPCR) analysis in Bos taurus semen.
  • To assess the expression stability of candidate reference miRNAs under varying semen quality conditions (motility and morphology).
  • To provide reliable normalizers for accurate miRNA expression studies in cattle reproduction.

Main Methods:

  • Selection of candidate reference small RNAs, including U6 small nuclear RNA and six specific miRNAs.
  • Analysis of miRNA expression stability using delta Ct, geNorm, NormFinder, and BestKeeper algorithms.
  • Comprehensive stability assessment using RefFinder for individual and combined semen quality conditions.

Main Results:

  • miR-92a-3p demonstrated the highest stability for normalizing analyses related to sperm motility.
  • Let-7c-5p was identified as the most stable reference miRNA for morphology-focused analyses.
  • Let-7c-5p was also the optimal normalizer for analyzing samples concurrently, while miR-26a-5p showed the least stability.

Conclusions:

  • This study provides the first validated set of microRNA (miRNA) normalizers for quantitative polymerase chain reaction (qPCR) in Bos taurus semen.
  • The identified normalizers (miR-92a-3p for motility, Let-7c-5p for morphology) enable more accurate miRNA quantification in cattle reproductive studies.
  • These findings will enhance the understanding of miRNA's role in cattle fertility and improve breeding program outcomes.