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Transcription Analysis of Clustered Essential Genes

ABSTRACT: Transcription Analysis of Clustered Essential Genes

PROVIDER: PRJNA612782 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR11310268.fastq.gz	Fastqsanger.gz
	SRR11310269.fastq.gz	Fastqsanger.gz
	SRR11310270.fastq.gz	Fastqsanger.gz
	SRR11310271.fastq.gz	Fastqsanger.gz
	SRR11310272.fastq.gz	Fastqsanger.gz

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Transcription Analysis of Clustered Essential Genes

Project description:All 14 essential genes of chrIII were clustered and reordered, and transcription of the genes was tested. We designed and constructed three types of reorganized essential genes into three different plasmids. For the genomic direction (GD) plasmid, the essential genes were arranged according to their natural chromosomal positions and orientations. For the same direction (SD) plasmid, the essential genes were arrayed in the same positions as GD plasmid, but all genes were assembled in the same direction (SD). For the random direction (RD) plasmid, the essential genes were modularly assembled according to the combined length of three individual genes and thus displayed disorderly. The synIII strains bearing the GD/SD/RD plasmids were phenotypically normal despite of the double expression of 14 essential genes.

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Project description:The maintenance of pluripotency requires the coordinated expression of a set of essential genes. Thorough identification of these genes is vital for gaining a better understanding of the pluripotent state. To this end, we have recently generated a genome-wide loss-of-function library in haploid human pluripotent stem cells utilizing the bacterial clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology using nearly 180,000 single guide RNAs (sgRNAs) and targeting 18,166 protein-coding genes. By combining the results of our high-throughput genetic screening with the analysis of publicly available RNAseq data of pluripotent stem cells and several somatic cells, we have determined the essential genes which have an enriched expression in hESCs and termed these genes as hESC-essential genes. Here, we have individually knocked down four of these hESC-essential genes with siRNAs, namely SALL4, DSCC1, SEPHS1 and VRTN, in human embryonic stem cells and analyzed the transcriptomes of the cells upon downregulation of these transcripts in order to unravel part of the pathways affected by hESC-essential genes.

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