Selecting Normalizers for MicroRNA RT-qPCR Expression Analysis in Murine Preimplantation Embryos and the Associated Conditioned Culture Media.

Normalizing RT-qPCR miRNA datasets that encompass numerous preimplantation embryo stages requires the identification of miRNAs that may be used as stable reference genes. A need has also arisen for the normalization of the accompanying conditioned culture media as extracellular miRNAs may serve as biomarkers of embryo developmental competence. Here, we evaluate the stability of six commonly used miRNA normalization candidates, as well as small nuclear U6, using five different means of evaluation (BestKeeper, NormFinder, geNorm, the comparative Delta Ct method and RefFinder comprehensive analysis) to assess their stability throughout murine preimplantation embryo development from the oocyte to the late blastocyst stages, both in whole embryos and the associated conditioned culture media.

Murine Blastocysts Release Mature MicroRNAs Into Culture Media That Reflect Developmental Status.

Extracellular microRNA (miRNA) sequences derived from the pre-implantation embryo have attracted interest for their possible contributions to the ongoing embryonic-uterine milieu, as well as their potential for use as accessible biomarkers indicative of embryonic health. Spent culture media microdroplets used to culture late-stage E4.0 murine blastocysts were screened for 641 mature miRNA sequences using a reverse transcription-quantitative polymerase chain reaction-based array.

Extracellular vesicles, microRNA and the preimplantation embryo: non-invasive clues of embryo well-being.

Elective single embryo transfer is rapidly becoming the standard of care in assisted reproductive technology for patients under the age of 35 years with a good prognosis. Clinical pregnancy rates have become increasingly dependent on the selection of a single viable embryo for transfer, and diagnostic techniques facilitating this selection continue to develop. Current progress in elucidating the extracellular vesicle and microRNA components of the embryonic secretome is reviewed, and the potential for these findings to improve clinical embryo selection discussed.