Methods and compositions for modulating a target genome are disclosed.

Methods and compositions for modulating a target genome are disclosed.

The present invention relates to a method for preparing probes for target nucleic acids targets. This method includes: a) obtaining a target DNA sequence of interest; b) adding adapter sequences to both ends of a fragmented DNA sequence while fragmenting the target DNA sequence by using transposase; and c) obtaining the fragmented DNA sequence by using the adapter sequences to generate the probes. The method provided by the present invention can efficiently, easily, and accurately mark the position of the genome at the level of one kilobase resolution.

The present invention relates to a method for preparing probes for target nucleic acids targets. This method includes: a) obtaining a target DNA sequence of interest; b) adding adapter sequences to both ends of a fragmented DNA sequence while fragmenting the target DNA sequence by using transposase; and c) obtaining the fragmented DNA sequence by using the adapter sequences to generate the probes. The method provided by the present invention can efficiently, easily, and accurately mark the position of the genome at the level of one kilobase resolution.

The present disclosure provides methods and systems for resolving cointegrate products from RNA-guided DNA integration. More particularly, the present disclosure provides systems comprising: an engineered Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-CRISPR associated (Cas) (CRISPR-Cas) system, and/or one or more vectors encoding the engineered CRISPR-Cas system, wherein the CRISPR-Cas system comprises: (a) at least one Cas protein, and (b) a guide RNA (gRNA); an engineered transposon system, and/or one or more vectors encoding the engineered transposon system, wherein the transposon system is configured for replicative transposition; a recombinase, or catalytic domain thereof, and/or one or more vectors encoding a recombinase, or catalytic domain thereof; and at least one donor nucleic acid to be integrated, wherein the donor nucleic acid comprises a recognition site for the recombinase and a cargo nucleic acid flanked by at least one transposon end sequence, and methods of use thereof.

The present disclosure provides methods and systems for resolving cointegrate products from RNA-guided DNA integration. More particularly, the present disclosure provides systems comprising: an engineered Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-CRISPR associated (Cas) (CRISPR-Cas) system, and/or one or more vectors encoding the engineered CRISPR-Cas system, wherein the CRISPR-Cas system comprises: (a) at least one Cas protein, and (b) a guide RNA (gRNA); an engineered transposon system, and/or one or more vectors encoding the engineered transposon system, wherein the transposon system is configured for replicative transposition; a recombinase, or catalytic domain thereof, and/or one or more vectors encoding a recombinase, or catalytic domain thereof; and at least one donor nucleic acid to be integrated, wherein the donor nucleic acid comprises a recognition site for the recombinase and a cargo nucleic acid flanked by at least one transposon end sequence, and methods of use thereof.

There is provided a DNA amplification method, comprising: preparation of a reaction solution containing a template DNA comprising a microsatellite, primers, a polymerase, and a recombinase, and subjection of the reaction solution to constant temperature incubation so as to amplify a DNA sequence comprising the microsatellite in the template DNA.

There is provided a DNA amplification method, comprising: preparation of a reaction solution containing a template DNA comprising a microsatellite, primers, a polymerase, and a recombinase, and subjection of the reaction solution to constant temperature incubation so as to amplify a DNA sequence comprising the microsatellite in the template DNA.

The present disclosure provides methods, systems, and kits for processing nucleic acid molecules. A method may comprise providing a template nucleic acid fragment (e.g., within a cell, cell bead, or cell nucleus) within a partition (e.g., a droplet or well) and subjecting the template nucleic acid fragment to one or more processes including a barcoding process and a single primer extension or amplification process. The processed template nucleic acid fragment may then be recovered from the partition and subjected to further amplification to provide material for subsequent sequencing analysis. The methods provided herein may permit simultaneous processing and analysis of both DNA and RNA molecules originating from the same cell, cell bead, or cell nucleus.

The present disclosure provides methods, systems, and kits for processing nucleic acid molecules. A method may comprise providing a template nucleic acid fragment (e.g., within a cell, cell bead, or cell nucleus) within a partition (e.g., a droplet or well) and subjecting the template nucleic acid fragment to one or more processes including a barcoding process and a single primer extension or amplification process. The processed template nucleic acid fragment may then be recovered from the partition and subjected to further amplification to provide material for subsequent sequencing analysis. The methods provided herein may permit simultaneous processing and analysis of both DNA and RNA molecules originating from the same cell, cell bead, or cell nucleus.