Illumina HiSeq 2000 sequencing; GSM3020753: WT_MII_A1 [libA.A1]; Mus musculus; RNA-Seq

WT_MII_A1 [libA.A1]

Pleomorphic adenoma gene 1 (PLAG1) encodes a transcription factor involved in cancer and growth. We study the role of PLAG1 in preimplantation embryos using STRT RNA-seq of single embryos from wild type and knockout mothers (both mated with wild type studs). The lack of maternal Plag1 led to delayed mouse 2-cell stage embryo development, compensatory expression of Plag1 from the paternal allele, and dysregulation of 1,089 genes. Half of these genes displayed a pattern of delayed activation and play roles in ribosome biogenesis and protein synthesis. These mouse genes further showed a significant overlap with human EGA genes with similar ontology, and an enrichment of the PLAG1 de novo motif. We conclude that Plag1 affects EGA through retrotransposons influencing ribosomes and protein synthesis, a mechanism that might also explain its roles in cancer and growth Overall design: Single wild type and maternal Plag1 knockout embryos at MII, 2-cell and 8-cell stage development in 14-16 biologicla replicas per developmental stage and genotype.

Maternal Plag1 deficiency delays two-cell stage embryo development and embryonic genome activation [Embryos]

Submitted by Gene Expression Omnibus on 22-JUN-2019

No extraction. Single embryos or uncultured oocytes were picked into 4 ml library lysis buffer containing 5 mM Tris-HCl pH 8.0 (Sigma-Aldrich), 2 mM dNTP mixture (ThermoFisher Scientific, Waltham, MA, USA), 10 mM DTT (Sigma-Aldrich), 0.05% Triton X-100 (Sigma-Aldrich), 400 nM anchored oligo(dT) primer biotin-TTAAGCAGTGGTATCAACGCAGAGTCGAC(T)29V where V is LNA nucleotide (Exiqon, Vedbaek, Denmark), and 4 U RiboLock RNase inhibitor (ThermoFisher). Two separate 46-plex libraries (libA and libB) were made as described previously (Krjutskov et al. 2016 Hum Reprod 31:844) with the following modifications: barcoded 10 µM template-switching oligonucleotides were added prior to reverse transcriptase, ERCC spike-in Mix A was diluted 15,300-fold with clean water, and 1 µl was taken per library reverse transcriptase master mix. Twenty cycles of PCR were used for the first round of amplification and ten additional cycles for the second round to introduce Illumina-compatible universal sequences. Both libraries contained all developmental stages and genotypes.

Maternal Plag1 deficiency delays two-cell stage embryo development and embryonic genome activation [Embryos]

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