Directed Differentiation of Trophoblast Stem Cells Provides a Useful In Vitro Model of Placenta Development in Cattle [RNA-Seq]

By utilizing a previously established bovine trophoblast stem cell (bTSC) model, we developed an optimized differentiation strategy to capture the complete developmental trajectory of placental binucleate cells (BNCs). This study provides new insights, demonstrating that extending the culture duration allows bTSCs to overcome previous developmental bottlenecks and faithfully recapitulate the in vivo BNC program, including the expression of late-stage secretory markers. Furthermore, we have shown that this tractable in vitro system serves as a robust platform for functional genomics, as evidenced by our validation of the transcription factor GCM1 as a driver of BNC differentiation. Overall design: The trophoblast cells from undifferentiated, day 4 of differentiation and day 10 of differentiation were collected and RNA was extracted using RNAsy microkit and sequenced by Azenta Life Sciences using three replicates each time point. Libraries were prepared using the NEBNext Ultra II RNA Library Prep for Illumina and sequenced (2 x 150 bp paired end) using Illumina instrument (4000 or equivalent). Raw sequence data (.bcl files) generated from Illumina Hiseq was converted into fastq files and de-multiplexed using Illumina's bcl2fastq 2.17 software.