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Highly efficient prime editing by introducing same-sense mutations in pegRNA or stabilizing its structure.

ABSTRACT: Highly efficient prime editing by introducing same-sense mutations in pegRNA or stabilizing its structure.

PROVIDER: PRJNA811688 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR18187913_1.fastq.gz	Fastqsanger.gz
	SRR18187913_2.fastq.gz	Fastqsanger.gz
	SRR18187914_1.fastq.gz	Fastqsanger.gz
	SRR18187914_2.fastq.gz	Fastqsanger.gz
	SRR18187915_1.fastq.gz	Fastqsanger.gz

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Highly efficient prime editing by introducing same-sense mutations in pegRNA or stabilizing its structure.

Project description:Prime editors (PEs) can mediate versatile genome editing but their efficiency remains low. Here, we developed spegRNA by introducing same-sense mutations at proper positions in the reverse-transcription template of pegRNA to increase PE’s single-base editing efficiency or apegRNA by altering the pegRNA secondary structure to increase PE’s indel-editing efficiency . When used in PE3 and PE5, the efficiencies of sPE3, aPE3, sPE5 and aPE5 were all enhanced significantly.

2022-03-10 | GSE197730 | GEO

Highly efficient prime editing by introducing same-sense mutations in pegRNA or stabilizing its structure.

Project description:Prime editor (PE), which is developed by combining Cas9 nickase and an engineered reverse transcriptase, can mediate all twelve types of base substitutions and small insertions or deletions in living cells but its efficiency remains low. Here, we develop spegRNA by introducing same-sense mutations at proper positions in the reverse-transcription template of pegRNA to increase PE's base-editing efficiency up-to 4,976-fold (on-average 353-fold). We also develop apegRNA by altering the pegRNA secondary structure to increase PE's indel-editing efficiency up-to 10.6-fold (on-average 2.77-fold). The spegRNA and apegRNA can be combined to further enhance editing efficiency. When spegRNA and apegRNA are used in PE3 and PE5 systems, the efficiencies of sPE3, aPE3, sPE5 and aPE5 systems are all enhanced significantly. The strategies developed in this study realize highly efficient prime editing at certain previously uneditable sites.

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Project description:Increasing numbers of sense–antisense transcripts (SATs), which are transcribed from the same chromosomal location but in opposite directions, have been identified in various eukaryotic species, but the biological meanings of most SATs remain unclear. To improve understanding of natural sense–antisense transcription, we performed comparative expression profiling of SATs conserved among humans and mice. Using custom oligo-arrays loaded with probes that represented SATs with both protein-coding and non-protein–coding transcripts, we showed that 33% of the 291 conserved SATs displayed identical expression patterns in the two species. Among these SATs, expressional balance inversion of sense–antisense genes was mostly observed in testis at a tissue-specific manner. Northern analyses of the individual conserved SAT loci revealed that: (1) a smeary hybridization pattern was present in mice, but not in humans, and (2) small RNAs (about 60 to 80 nt) were detected from the exon-overlapping regions of SAT loci. In addition, further analyses showed marked alteration of sense–antisense expression balance throughout spermatogenesis in testis. These results suggest that conserved SAT loci are rich in potential regulatory roles that will help us understand this new class of transcripts underlying the mammalian genome. Keywords: Expression profile of mouse and human sense-antisense transcript

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