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Preparation of Small RNA Libraries for Sequencing from Early Mouse Embryos
Published on: October 9, 2020
Differential expression of microRNAs in 2-cell and 4-cell mouse embryos
Pei Wang1, Ji Cui1, Chun Zhao1
1State Key Laboratory of Reproductive Medicine,Department of Reproduction,Nanjing Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University,Nanjing 210004,China.
Abstract:
In vitro fertilized (IVF) human embryos have a high incidence of developmental arrest before the blastocyst stage, therefore characterization of the molecular mechanisms that regulate embryo development is urgently required. Post-transcriptional control by microRNAs (miRNAs) is one of the most investigated RNA control mechanisms, and is hypothesized to be involved actively in developmental arrest in preimplantation embryos. In this study, we extracted total RNA from mouse 2-cell and 4-cell embryos. Using a miRNA microarray, 192 miRNAs were found to be differentially expressed in 4-cell embryos and 2-cell embryos; 122 miRNAs were upregulated and 70 were downregulated in 4-cell embryos. The microarray results were confirmed by real-time quantitative RT-PCR for six miRNAs (mmu-miR-467h, mmu-miR-466d-3p, mmu-miR-292-5p, mmu-miR-154, mmu-miR-2145, and mmu-miR-706). Cdca4 and Tcf12 were identified as miR-154 target genes by target prediction analysis. This study provides a developmental map for a large number of miRNAs in 2-cell and 4-cell embryos. The function of these miRNAs and the mechanisms by which they modulate embryonic developmental arrest require further study. The results of this study have potential applications in the field of reproductive medicine.
Insights
MicroRNAs (miRNAs) play a role in early embryo development. This study mapped differentially expressed miRNAs in mouse embryos, identifying potential targets involved in developmental regulation.
Area of Science:
- Developmental Biology
- Molecular Biology
- Reproductive Medicine
Background:
- In vitro fertilized (IVF) human embryos frequently arrest development before the blastocyst stage.
- Understanding molecular regulation of early embryo development is crucial.
- Post-transcriptional regulation by microRNAs (miRNAs) is a key mechanism.
Purpose of the Study:
- To characterize differentially expressed miRNAs in mouse 2-cell and 4-cell embryos.
- To identify potential miRNA targets involved in embryonic development.
- To provide a miRNA developmental map for early mouse embryos.
Main Methods:
- Total RNA extraction from mouse 2-cell and 4-cell embryos.
- miRNA microarray analysis to identify differentially expressed miRNAs.
- Real-time quantitative RT-PCR validation for selected miRNAs.
- Bioinformatic target prediction analysis for identified miRNAs.
Main Results:
- 192 miRNAs were differentially expressed between 2-cell and 4-cell mouse embryos.
- 122 miRNAs were upregulated and 70 downregulated in 4-cell embryos.
- Six specific miRNAs were validated using RT-PCR.
- Cdca4 and Tcf12 were predicted as target genes for mmu-miR-154.
Conclusions:
- This study presents a comprehensive miRNA expression profile during early mouse embryonic development.
- The identified miRNAs and their predicted targets offer insights into mechanisms regulating embryonic development.
- Further research is needed to elucidate miRNA functions in preventing embryonic developmental arrest.
- Findings have potential implications for improving reproductive medicine outcomes.
