The present invention relates to, among other things, probes, compositions, methods, and kits for simultaneous, multiplexed detection and quantification of protein and/or nucleic acid expression in a user-defined region of a tissue, user-defined cell, and/or user-defined subcellular structure within a cell.

The present invention provides a use of a Cas protein, and a method and a kit for detecting target nucleic acid molecules. The method for detecting target nucleic acid molecules comprises adding a guide RNA, a Cas12a, and a nucleic acid probe into a reaction system containing target nucleic acid molecules to be detected, and detecting it after the reaction is completed.

The present invention provides a use of a Cas protein, and a method and a kit for detecting target nucleic acid molecules. The method for detecting target nucleic acid molecules comprises adding a guide RNA, a Cas12a, and a nucleic acid probe into a reaction system containing target nucleic acid molecules to be detected, and detecting it after the reaction is completed.

The present invention provides a use of a Cas protein, and a method and a kit for detecting target nucleic acid molecules. The method for detecting target nucleic acid molecules comprises adding a guide RNA, a Cas12a, and a nucleic acid probe into a reaction system containing target nucleic acid molecules to be detected, and detecting it after the reaction is completed.

The present disclosure describes compositions of matter comprising a ribonucleoprotein complex comprising a nucleic acid-guided nuclease and a guide RNA, and further comprising and a blocking nucleic acid molecule represented by Formula I, wherein Formula I in the 5′-to-3′ direction comprises: A-(B-L).sub.J-C-M-T-D; wherein A is 0-15 nucleotides in length; B is 4-12 nucleotides in length; L is 3-25 nucleotides in length; J is an integer between 1 and 10; C is 4-15 nucleotides in length; M is 1-25 nucleotides in length or is absent, wherein if M is absent then A-(B-L).sub.J-C and T-D are separate nucleic acid strands; T is 17-135 nucleotides in length and comprises at least 50% sequence complementarity to B and C; D is 0-10 nucleotides in length and comprises at least 50% sequence complementarity to A; and wherein the blocking nucleic acid molecule comprises a sequence complementary to a gRNA.

The present disclosure describes compositions of matter comprising a ribonucleoprotein complex comprising a nucleic acid-guided nuclease and a guide RNA, and further comprising and a blocking nucleic acid molecule represented by Formula I, wherein Formula I in the 5′-to-3′ direction comprises: A-(B-L).sub.J-C-M-T-D; wherein A is 0-15 nucleotides in length; B is 4-12 nucleotides in length; L is 3-25 nucleotides in length; J is an integer between 1 and 10; C is 4-15 nucleotides in length; M is 1-25 nucleotides in length or is absent, wherein if M is absent then A-(B-L).sub.J-C and T-D are separate nucleic acid strands; T is 17-135 nucleotides in length and comprises at least 50% sequence complementarity to B and C; D is 0-10 nucleotides in length and comprises at least 50% sequence complementarity to A; and wherein the blocking nucleic acid molecule comprises a sequence complementary to a gRNA.

The present disclosure describes compositions of matter comprising a ribonucleoprotein complex comprising a nucleic acid-guided nuclease and a guide RNA, and further comprising and a blocking nucleic acid molecule represented by Formula I, wherein Formula I in the 5′-to-3′ direction comprises: A-(B-L).sub.J-C-M-T-D; wherein A is 0-15 nucleotides in length; B is 4-12 nucleotides in length; L is 3-25 nucleotides in length; J is an integer between 1 and 10; C is 4-15 nucleotides in length; M is 1-25 nucleotides in length or is absent, wherein if M is absent then A-(B-L).sub.J-C and T-D are separate nucleic acid strands; T is 17-135 nucleotides in length and comprises at least 50% sequence complementarity to B and C; D is 0-10 nucleotides in length and comprises at least 50% sequence complementarity to A; and wherein the blocking nucleic acid molecule comprises a sequence complementary to a gRNA.

The present disclosure describes compositions of matter comprising a ribonucleoprotein complex comprising a nucleic acid-guided nuclease and a guide RNA, and further comprising and a blocking nucleic acid molecule represented by Formula I, wherein Formula I in the 5′-to-3′ direction comprises: A-(B-L).sub.J-C-M-T-D; wherein A is 0-15 nucleotides in length; B is 4-12 nucleotides in length; L is 3-25 nucleotides in length; J is an integer between 1 and 10; C is 4-15 nucleotides in length; M is 1-25 nucleotides in length or is absent, wherein if M is absent then A-(B-L).sub.J-C and T-D are separate nucleic acid strands; T is 17-135 nucleotides in length and comprises at least 50% sequence complementarity to B and C; D is 0-10 nucleotides in length and comprises at least 50% sequence complementarity to A; and wherein the blocking nucleic acid molecule comprises a sequence complementary to a gRNA.

The disclosure describes novel systems, methods, and compositions for the manipulation of nucleic acids in a targeted fashion. The disclosure describes non-naturally occurring, engineered CRISPR systems, components, and methods for targeted modification of nucleic acids. Each system includes one or more protein components and one or more nucleic acid components that together target nucleic acids.

The disclosure describes novel systems, methods, and compositions for the manipulation of nucleic acids in a targeted fashion. The disclosure describes non-naturally occurring, engineered CRISPR systems, components, and methods for targeted modification of nucleic acids. Each system includes one or more protein components and one or more nucleic acid components that together target nucleic acids.