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Updated: Mar 29, 2026
Defining the Program of Maternal mRNA Translation during In vitro Maturation using a Single Oocyte Reporter Assay
Published on: June 16, 2021
Evaluation of ribonucleic acid amplification protocols for human oocyte transcriptome analysis.
Eleni Mantikou1, Oskar Bruning2, Sebastiaan Mastenbroek3
1Center for Reproductive Medicine, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands; MicroArray Department and Integrative Bioinformatics Unit, Swammerdam Institute for Life Sciences, Faculty of Science, University of Amsterdam, Amsterdam, the Netherlands.
Two RNA amplification protocols effectively profile gene expression in single human oocytes, providing reliable and reproducible results for research. These methods yield sufficient messenger RNA (mRNA) for various downstream applications, including sequencing.
Area of Science:
- Reproductive biology
- Molecular genetics
- Genomics
Background:
- Investigating gene expression in individual human oocytes is crucial for understanding reproductive health and disease.
- Existing methods for messenger RNA (mRNA) amplification from limited biological samples often lack reliability and reproducibility.
Purpose of the Study:
- To develop and validate a dependable method for analyzing gene-expression profiles from single human oocytes.
- To assess the efficiency and reproducibility of commercially available mRNA amplification protocols for oocyte samples.
Main Methods:
- Five commercial messenger RNA (mRNA) amplification protocols were tested on 54 single human oocytes.
- Protocols yielding sufficient mRNA were validated using microarray analysis on 25 oocytes.
- A validation strategy involved pooling and splitting RNA from 13 oocytes to assess technical variation.
Main Results:
- Two protocols, WT-Ovation One-Direct and Arcturus RiboAMp HS Plus, demonstrated sufficient yields and high success rates.
- Both validated protocols produced reliable, reproducible gene-expression profiles (r ≥ 0.95) with low technical variation.
- One protocol identified more transcripts but also yielded a higher rate of false discoveries.
Conclusions:
- Two tested protocols generate adequate mRNA for downstream applications like quantitative polymerase chain reaction, microarrays, and sequencing.
- Both protocols are suitable for gene-expression profiling of individual human oocytes, even when distinguishing biological from technical variation.
