Murine blastocysts release mature microRNAs into culture media that reflect developmental status

Extracellular microRNA sequences have attracted interest for their potential use as accessible biomarkers that may be non-invasively collected from a wide range of biological fluids, including spent culture media. The ongoing milieu between the late-stage preimplantation embryo and the receptive uterus remains an actively investigated area of research and the physiological role of secreted microRNA sequences remains to be determined. Spent culture media used to culture late stage E4.0 murine blastocysts was screened for 641 mature microRNA sequences using a qPCR-based array. We report here the 39 sequences that were exclusively detected in the conditioned media using this approach. The embryonic miR-290 family were among the most abundant extracellular sequences. Notably, implantation-relevant members miR-126a, miR-101a, miR-143 and miR-320 were also detected. Spent murine blastocyst culture was analyzed for mature microRNA using TaqMan Rodent MicroRNA Array v3.0 A + B Cards after RT using a TaqMan MicroRNA Reverse Transcription Kit with Megaplex RT Primers and preamplification using Megaplex PreAmp Primer Pools, Rodent Pools Set v3.0. 2 samples were analyzed: 1) a 6 µL sample of 8 µL culture media conditioned with 25 murine CD1 blastocysts from 99 h to 116 h post-hCG injection, 2) a negative control and contains an equivalent volume of blank culture media.