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SRX3049471: GSM2722410: endo1; Xenopus laevis; RNA-Seq
1 ILLUMINA (Illumina HiSeq 2500) run: 57.8M spots, 11.6G bases, 3.8Gb downloads

Submitted by: NCBI (GEO)
Study: The Xenopus Primordial Germ Cell Transcriptome: Unexpected Role for sox7 in Early PGC Development
show Abstracthide Abstract
Xenopus primordial germ cells (PGCs) are determined by the presence of maternally derived germ plasm. Germ plasm components both protect PGCs from somatic differentiation and begin a unique gene expression program. Segregation of the germline from the endodermal lineage occurs during gastrulation and PGCs subsequently initiate zygotic transcription. However, the gene-networks that operate to both preserve the potential for totipotency and promote germline differentiation are poorly understood. Here, we utilized RNA-sequencing analysis to comprehensively interrogate PGC and neighboring endoderm cell RNAs after lineage segregation. We identified 1,865 transcripts enriched in PGCs compared to endoderm cells. Over 50% of maternal, vegetally-enriched transcripts were enriched in the PGC transcriptome, including sox7. PGC-directed sox7 knockdown and over-expression studies revealed an early requirement for sox7 in proper germ plasm localization, zygotic transcription, and PGC number. We identified oct60 as the most highly expressed and enriched OCT3/4 homologue in PGCs. Lastly, we compared the Xenopus PGC transcriptome with human PGC transcripts and showed that 80% of transcripts are conserved, identifying Xenopus as a relevant model system for understanding the gene-networks necessary for human germline development. Overall design: Examination of X. laevis primordial germ cell (PGC) and neighboring endoderm cell (Endo) RNAs after lineage segregation to determine PGC-enriched transcripts that may contribute to germline development.
Sample: endo1
SAMN07427078 • SRS2396161 • All experiments • All runs
Organism: Xenopus laevis
Library:
Instrument: Illumina HiSeq 2500
Strategy: RNA-Seq
Source: TRANSCRIPTOMIC
Selection: cDNA
Layout: PAIRED
Construction protocol: Primordial germ cells and stage-matched endoderm cells were isolated from stage 12-14 Xenopus laevis embryos and collected into RNA lysis buffer as described by Butler et al. (2016). Total RNA was extracted using the RNAqueous-Micro Total RNA Isolation Kit per the manufacturer’s protocol (Ambion). Three 1ng aliquots of each rRNA-depleted sample were used as template for preparation of sequence-able template DNA molecules using the ScriptSeq v2 RNASeq Library Preparation Kit (Illumina, Part # SSV21124). The quality and size distribution of the amplified libraries were determined utilizing an Agilent 2100 Bioanalyzer High Sensitivity DNA Chip. Libraries were quantified using the KAPA Library Quantification Kit (Kapa Biosystems, Boston, MA), and equimolar concentrations were diluted prior to loading onto the flow cell of the Illumina cBot cluster station. The libraries were extended and bridge amplified to create sequence clusters using the Illumina HiSeq PE Cluster Kit v4, and sequenced on an Illumina HiSeq Flow Cell v4 with 100-bp paired-end reads plus index read using the Illumina HiSeq SBS Kit v4. Real time image analysis and base calling were performed on the instrument using the HiSeq Sequencing Control Software version 2.2.58. Three DNA libraries were prepared from Xenopus laevis primordial germ cell RNA and three from matched endoderm cell RNA.
Experiment attributes:
GEO Accession: GSM2722410
Links:
Runs: 1 run, 57.8M spots, 11.6G bases, 3.8Gb
Run# of Spots# of BasesSizePublished
SRR588338257,804,79011.6G3.8Gb2017-08-03

ID:
4328890

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