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Targeted site integration in plants using a transposon system

ABSTRACT: Targeted site integration in plants using a transposon system

PROVIDER: PRJNA943231 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR23802573.fastq.gz	Fastqsanger.gz
	SRR23802574.fastq.gz	Fastqsanger.gz
	SRR23802575.fastq.gz	Fastqsanger.gz
	SRR23802576.fastq.gz	Fastqsanger.gz
	SRR23802577.fastq.gz	Fastqsanger.gz

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Targeted site integration in plants using a transposon system [Amplicon]

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Targeted site integration in plants using a transposon system

Project description:This SuperSeries is composed of the SubSeries listed below.

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The Transposon Insertion site Profiling Chip (TIP-chip)

Project description:We have developed a microarray intended for use in finding all transposons in a region of interest. By selectively amplifying and hybridizing transposon flanking DNA to our array, we can localize all transposons in the region present on our TIP-chip, a dense tiling array. We have tested our technology in yeast and have been successful. Keywords: transposon insertion profiling, genomic DNA, yeast

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Project description:The CRISPR-Cas12a platform has attracted interest in the genome editing community because the prototypic Acidaminococcus Cas12a generates a staggered DNA double-strand break upon binding to an AT-rich protospacer-adjacent motif (PAM, 5'-TTTV). The broad application of the platform in primary human cells was enabled by the development of an engineered version of the natural Cas12a protein, called Cas12a Ultra. In this study, we confirmed that CRISPR-Cas12a Ultra ribonucleoprotein complexes enabled allelic gene disruption frequencies of over 90% at multiple target sites in human T cells, hematopoietic stem and progenitor cells (HSPCs), and induced pluripotent stem cells (iPSCs). In addition, we demonstrated for the first time the efficient knock-in potential of the platform in human iPSCs and achieved targeted integration of a GFP marker gene into the AAVS1 safe harbor site and a CSF2RA super-exon into CSF2RA in up to 90% of alleles without selection. Clonal analysis revealed bi-allelic integration in >50% of the screened iPSC clones without compromising their pluripotency and genomic integrity. Thus, in combination with the adeno-associated virus vector system, CRISPR-Cas12a Ultra provides a highly efficient genome editing platform for performing targeted knock-ins in human iPSCs.

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