Genomic editing of improved efficiency and accuracy
AssigneeShanghaitech University
InventorsJia CHEN, Bei YANG, Li YANG, Xiaosa LI, Xiao WANG, Wenwen ZHAO, Lina ZHOU, Jifang LI
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Abstract
Provided are compositions and methods for enhanced prime editing, which include a pegRNA that encodes a target mutation in a target protein, along with one or more nearby silent or conservative mutations. These silent mutations can increase the editing efficiency, without causing a change to the target protein sequence. Also provided are compositions and methods of using the improved prime editing for preventing or treating infections by SARS-CoV or SARS-CoV-2.
Claims20
A method for generating a target mutation to a protein in a cell, comprising introducing to the cell a prime editing system, wherein the prime editing system comprises a fusion protein comprising a nickase and a reverse transcriptase, and a prime editing guide RNA (pegRNA) encoding the target mutation and one or more silent or conservative mutations within 20 nucleotides from the target mutation.
A method for introducing mutations to a protein in a cell, comprising introducing to the cell a prime editing system, wherein the prime editing system comprises a fusion protein comprising a nickase and a reverse transcriptase, and a prime editing guide RNA (pegRNA) encoding two or more mutations within the coding sequence of the protein, wherein at least one of the mutations is a silent or conservative mutation and is within 20 nucleotides from another of mutations.
14-29. (canceled)
A method for conducting genetic editing in a cell at a target site, comprising introducing to the cell:
- a first construct encoding a nickase, and
- one or more second construct encoding (a) a prime editing guide RNA (pegRNA) capable of identifying the target site and including genetic information for editing the target site, (b) a single guide RNA (sgRNA) capable of directing the nickase to nick a non-edited DNA strand of the target site, wherein the pegRNA or the sgRNA includes a tag sequence, and (c) a reverse-transcriptase fused to an RNA recognition peptide capable of binding to the tag sequence.
Dependencies are inferred from the claim text.