Aspects of the present disclosure relate generally to methods, compositions, and kits for in situ whole cell barcoding. Aspects of the present disclosure also include a computer readable-medium and a processor to carry out the steps of the method described herein. In some embodiments, the disclosure relates to whole cell barcoding performed in situ.

Aspects of the present disclosure relate generally to methods, compositions, and kits for in situ whole cell barcoding. Aspects of the present disclosure also include a computer readable-medium and a processor to carry out the steps of the method described herein. In some embodiments, the disclosure relates to whole cell barcoding performed in situ.

The present invention relates to a method for detecting an integration pattern of a virus in a host genome. In particular, a method is provided encompassing selective cleavage of circularized DNA fragments carrying viral DNA with an RNA-guided endonuclease and at least one guide RNA or at least one pool of guide RNAs, followed by inverse PCR, in particular inverse long-range PCR, and sequencing. The invention further relates to kits for performing the method and application of the method.

The present invention relates to a method for detecting an integration pattern of a virus in a host genome. In particular, a method is provided encompassing selective cleavage of circularized DNA fragments carrying viral DNA with an RNA-guided endonuclease and at least one guide RNA or at least one pool of guide RNAs, followed by inverse PCR, in particular inverse long-range PCR, and sequencing. The invention further relates to kits for performing the method and application of the method.

The present invention relates to the field of ribonucleotide analysis. More specifically, the present invention provides compositions and methods for detection for nucleic acids using probe pair litigation. In particular embodiments, the compositions and methods of the present invention utilize a probe set comprising (1) a first multi-partite probe comprising a 5′ phosphorylated donor probe and a first bridge probe, wherein the 5′ phosphorylated donor probe specifically hybridizes to a target nucleic acid; and (ii) a second multi-partite probe comprising a 3′ acceptor probe and a second bridge probe, wherein the 3′ acceptor probe specifically hybridizes to the target nucleic acid adjacent to the 5′ donor probe and the second bridge probe is 5′ phosphorylated.

The present invention relates to the field of ribonucleotide analysis. More specifically, the present invention provides compositions and methods for detection for nucleic acids using probe pair litigation. In particular embodiments, the compositions and methods of the present invention utilize a probe set comprising (1) a first multi-partite probe comprising a 5′ phosphorylated donor probe and a first bridge probe, wherein the 5′ phosphorylated donor probe specifically hybridizes to a target nucleic acid; and (ii) a second multi-partite probe comprising a 3′ acceptor probe and a second bridge probe, wherein the 3′ acceptor probe specifically hybridizes to the target nucleic acid adjacent to the 5′ donor probe and the second bridge probe is 5′ phosphorylated.

CRISPR RNA-guided nucleases are routinely used for sequence-specific manipulation of DNA. While CRISPR-based DNA editing has become routine, analogous methods for editing RNA have yet to be established. Here we repurpose the type III-A CRISPR RNA-guided nuclease for sequence-specific cleavage of the SARS-CoV-2 genome. The type III cleavage reaction is performed in vitro using purified viral RNA, resulting in sequence-specific excision of 6, 12, 18 or 24 nucleotides. Ligation of the cleavage products is facilitated by a DNA splint that bridges the excision and RNA ligase is used to link the RNA products before transfection into mammalian cells. The SARS-CoV-2 RNA is infectious and standard plaque assays are used to recover viral clones. Collectively, this work demonstrates how type III CRISPR systems can be repurposed for sequence-specific editing of RNA viruses including SARS-CoV-2 and more generally for gene therapy.

CRISPR RNA-guided nucleases are routinely used for sequence-specific manipulation of DNA. While CRISPR-based DNA editing has become routine, analogous methods for editing RNA have yet to be established. Here we repurpose the type III-A CRISPR RNA-guided nuclease for sequence-specific cleavage of the SARS-CoV-2 genome. The type III cleavage reaction is performed in vitro using purified viral RNA, resulting in sequence-specific excision of 6, 12, 18 or 24 nucleotides. Ligation of the cleavage products is facilitated by a DNA splint that bridges the excision and RNA ligase is used to link the RNA products before transfection into mammalian cells. The SARS-CoV-2 RNA is infectious and standard plaque assays are used to recover viral clones. Collectively, this work demonstrates how type III CRISPR systems can be repurposed for sequence-specific editing of RNA viruses including SARS-CoV-2 and more generally for gene therapy.

The present disclosure relates to a method of assembling long nucleic acids by enzymatically ligating oligonucleotide molecules hybridized to an indexed splint oligonucleotide molecules. Also disclosed are oligonucleotide structures comprising an indexed splint oligonucleotide useful in performing the disclosed method.

The present disclosure relates to a method of assembling long nucleic acids by enzymatically ligating oligonucleotide molecules hybridized to an indexed splint oligonucleotide molecules. Also disclosed are oligonucleotide structures comprising an indexed splint oligonucleotide useful in performing the disclosed method.