Proximity-mediated rolling circle amplification
First Claim
1. A method comprising:
- (a) bringing into contact one or more analyte samples and one or more binding guide conjugates, wherein each binding guide conjugate comprises a specific binding molecule and a guide oligonucleotide, wherein each specific binding molecule interacts directly or indirectly with an analyte in the analyte sample, and incubating the analyte samples and the binding guide conjugates under conditions that promote interaction of the specific binding molecules and the analytes, (b) prior to, simultaneous with, or following step (a), bringing into contact the binding guide conjugates and one or more half circle probes, wherein each half circle probe comprises a single-stranded DNA molecule comprising two guide complement portions, wherein each guide complement portion is complementary to one of the guide oligonucleotides, and incubating the binding guide conjugates and the half circle probes under conditions that promote hybridization between the guide oligonucleotides and the half circle probes, (c) following step (b) and prior to, simultaneous with, or following step (a), incubating the binding guide conjugates and half circle probes under conditions that promote ligation of half circle probes, thereby producing one or more amplification target circles.
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Abstract
Disclosed are compositions and methods for proximity-mediated rolling circle amplification and for real-time detection of proximity-mediated rolling circle amplification products. Rolling circle amplification (RCA) refers to nucleic acid amplification reactions involving replication of a circular nucleic acid template to form a long strand with tandem repeats of the sequence complementary to the circular template. In proximity-mediated RCA, binding guide conjugates are brought into close proximity, generally by associating them to the same analyte or to two analytes in close proximity. The binding guide conjugates comprise a specific binding molecule and a guide oligonucleotide. The guide oligonucleotides are complementary to guide complement portions on half circle probes. The complementary sequences between the guide oligonucleotides and half circle probes allow both ends of each half circle probe to hybridize adjacent to an end of the other half circle probe and to be ligated together to form a circular nucleic acid molecule comprising the two half circle probes. This circular nucleic acid molecule can then be used as the template in RCA.
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Citations
138 Claims
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1. A method comprising:
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(a) bringing into contact one or more analyte samples and one or more binding guide conjugates, wherein each binding guide conjugate comprises a specific binding molecule and a guide oligonucleotide, wherein each specific binding molecule interacts directly or indirectly with an analyte in the analyte sample, and incubating the analyte samples and the binding guide conjugates under conditions that promote interaction of the specific binding molecules and the analytes, (b) prior to, simultaneous with, or following step (a), bringing into contact the binding guide conjugates and one or more half circle probes, wherein each half circle probe comprises a single-stranded DNA molecule comprising two guide complement portions, wherein each guide complement portion is complementary to one of the guide oligonucleotides, and incubating the binding guide conjugates and the half circle probes under conditions that promote hybridization between the guide oligonucleotides and the half circle probes, (c) following step (b) and prior to, simultaneous with, or following step (a), incubating the binding guide conjugates and half circle probes under conditions that promote ligation of half circle probes, thereby producing one or more amplification target circles. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94)
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95. A method comprising:
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(a) bringing into contact one or more analyte samples and one or more arrays, wherein each array comprises a set of analyte capture agents, a set of accessory molecules, or both, wherein each analyte capture agent interacts with an analyte directly or indirectly, (b) prior to, simultaneous with, or following step (a), bringing into contact one or more of the arrays and one or more binding guide conjugates, wherein each binding guide conjugate comprises a specific binding molecule and a guide oligonucleotide, wherein each specific binding molecule interacts directly or indirectly with an analyte in the analyte sample, and incubating the analyte samples and the binding guide conjugates under conditions that promote interaction of the specific binding molecules and the analytes, (c) prior to, simultaneous with, or following steps (a) or (b), bringing into contact the binding guide conjugates and one or more half circle probes, wherein each half circle probe comprises a single-stranded DNA molecule comprising two guide complement portions, wherein each guide complement portion is complementary to at least one of the guide oligonucleotides, and incubating the binding guide conjugates and the half circle probes under conditions that promote hybridization between the guide oligonucleotides and the half circle probes, (d) following step (c) and prior to, simultaneous with, or following steps (a) or (b), incubating the binding guide conjugates and half circle probes under conditions that promote ligation of half circle probes to each other, thereby producing one or more amplification target circles, and (e) following step (d), incubating the amplification target circles under conditions that promote replication of the amplification target circles, wherein replication of the amplification target circles results in the formation of tandem sequence DNA. - View Dependent Claims (96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117)
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118. A method comprising:
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(a) treating one or more analyte samples so that one or more analytes are modified, (b) prior to, simultaneous with, or following step (a), bringing into contact one or more of the analyte samples and one or more binding guide conjugates, wherein each binding guide conjugate comprises a specific binding molecule and a guide oligonucleotide, wherein each specific binding molecule interacts directly or indirectly with an analyte in the analyte sample, and incubating the analyte samples and the binding guide conjugates under conditions that promote interaction of the specific binding molecules and the analytes, (c) prior to, simultaneous with, or following steps (a) or (b), bringing into contact the binding guide conjugates and one or more half circle probes, wherein each half circle probe comprises a single-stranded DNA molecule comprising two guide complement portions, wherein each guide complement portion is complementary to at least one of the guide oligonucleotides, and incubating the binding guide conjugates and the half circle probes under conditions that promote hybridization between the guide oligonucleotides and the half circle probes, (d) following step (c) and prior to, simultaneous with, or following steps (a) or (b), incubating the binding guide conjugates and half circle probes under conditions that promote ligation of half circle probes to each other, thereby producing one or more amplification target circles, and (e) following step (d), incubating the amplification target circles under conditions that promote replication of the amplification target circles, wherein replication of the amplification target circles results in the formation of tandem sequence DNA. - View Dependent Claims (119)
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120. A method comprising:
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(a) bringing into contact one or more analyte samples and one or more arrays, wherein each array comprises a set of analyte capture agents, a set of accessory molecules, or both, wherein each analyte capture agent interacts with an analyte directly or indirectly, (b) prior to, simultaneous with, or following step (a), bringing into contact one or more of the analyte samples and one or more binding guide conjugates, wherein each binding guide conjugate comprises a specific binding molecule and a guide oligonucleotide, wherein each specific binding molecule interacts directly or indirectly with an analyte in the analyte sample, and incubating the analyte samples and the binding guide conjugates under conditions that promote interaction of the specific binding molecules and the analytes, (c) prior to, simultaneous with, or following steps (a) or (b), bringing into contact the binding guide conjugates and one or more half circle probes, wherein each half circle probe comprises a single-stranded DNA molecule comprising two guide complement portions, wherein each guide complement portion is complementary to at least one of the guide oligonucleotides, and incubating the binding guide conjugates and the half circle probes under conditions that promote hybridization between the guide oligonucleotides and the half circle probes, (d) following step (c) and prior to, simultaneous with, or following steps (a) or (b), incubating the binding guide conjugates and half circle probes under conditions that promote ligation of half circle probes to each other, thereby producing one or more amplification target circles, and (e) following step (d), incubating the amplification target circles under conditions that promote replication of the amplification target circles, wherein replication of the amplification target circles results in the formation of tandem sequence DNA.
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121. A kit comprising
(a) a plurality of binding guide conjugates, wherein each binding guide conjugates comprises a specific binding molecule and a guide oligonucleotide, wherein each specific binding molecule interacts with an analyte directly or indirectly, and (b) a plurality of half circle probes, wherein each half circle probe comprises a single-stranded DNA molecule comprising two guide complement portions, wherein each guide complement portion is complementary to at least one of the guide oligonucleotides.
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128. A method comprising:
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(a) bringing into contact one or more analyte samples and one or more binding half circle conjugates, wherein each binding half circle conjugate comprises a specific binding molecule and a half circle probe, wherein each specific binding molecule interacts directly or indirectly with an analyte in the analyte sample, and wherein each half circle probe comprises a single-stranded DNA molecule comprising two guide complement portions, and incubating the analyte samples and the binding half circle conjugates under conditions that promote interaction of the specific binding molecules and the analytes, (b) prior to, simultaneous with, or following step (a), bringing into contact the binding half circle conjugates and one or more guide oligonucleotides, wherein each guide oligonucleotide is complementary to at least one of the guide complement portions of the half circle probes, and incubating the binding half circle conjugates and the guide oligonucleotides under conditions that promote hybridization between the guide oligonucleotides and the half circle probes, (c) following step (b) and prior to, simultaneous with, or following step (a), incubating the binding half circle conjugates and guide oligonucleotides under conditions that promote ligation of half circle probes to each other, thereby producing one or more amplification target circles, and (d) following step (c), incubating the amplification target circles under conditions that promote replication of the amplification target circles, wherein replication of the amplification target circles results in the formation of tandem sequence DNA. - View Dependent Claims (129, 130, 131, 132, 133, 134, 135)
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136. A method comprising:
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(a) bringing into contact one or more analyte samples and one or more binding guide conjugates, wherein each binding guide conjugate comprises a specific binding molecule and a guide oligonucleotide, wherein each specific binding molecule interacts directly or indirectly with an analyte in the analyte sample, and incubating the analyte samples and the binding guide conjugates under conditions that promote interaction of the specific binding molecules and the analytes, (b) prior to, simultaneous with, or following step (a), bringing into contact the binding guide conjugates and one or more half circle probes, wherein each half circle probe comprises a single-stranded DNA molecule comprising two guide complement portions, wherein each guide complement portion is complementary to at least one of the guide oligonucleotides, and incubating the binding guide conjugates and the half circle probes under conditions that promote hybridization between the guide oligonucleotides and the half circle probes, (c) following step (b) and prior to, simultaneous with, or following step (a), incubating the binding guide conjugates and half circle probes under conditions that promote ligation of half circle probes to each other, thereby producing one or more amplification target circles, and (d) following step (c), incubating the binding guide conjugates and amplification target circles under conditions that promote replication of the amplification target circles, wherein replication of the amplification target circles results in the formation of tandem sequence DNA.
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137. A method comprising:
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(a) bringing into contact one or more analyte samples and one or more binding guide conjugates, wherein each binding guide conjugate comprises a specific binding molecule and a guide oligonucleotide, wherein each specific binding molecule interacts directly or indirectly with an analyte in the analyte sample, and incubating the analyte samples and the binding guide conjugates under conditions that promote interaction of the specific binding molecules and the analytes, (b) prior to, simultaneous with, or following step (a), bringing into contact the binding guide conjugates and one or more half circle probes, wherein each half circle probe comprises a single-stranded DNA molecule comprising two guide complement portions, wherein each guide complement portion is complementary to at least one of the guide oligonucleotides, wherein each guide oligonucleotide is complementary to one of the guide complement portions of both half circle probes in a pair of half circle probes, wherein the guide complement portions of the same half circle probe are complementary to two different guide oligonucleotides, wherein the two different guide oligonucleotides that each are complementary to a different one of the guide complement portions of the same half circle probe constitute a pair of guide oligonucleotides, wherein the binding guide conjugates that comprise the guide oligonucleotides in a pair of guide oligonucleotides constitute a pair of binding guide conjugates, wherein one guide complement portion of each half circle probe in a pair of half circle probes is complementary to one of the guide oligonucleotides in a pair of guide oligonucleotides and the other guide complement portion of each half circle probe in the pair of half circle probes is complementary to the other guide oligonucleotide in the pair of guide oligonucleotides, wherein both half circle probes in a pair of half circle probes are hybridized to both guide oligonucleotides in a pair of guide oligonucleotides, and incubating the binding guide conjugates and the half circle probes under conditions that promote hybridization between the guide oligonucleotides and the half circle probes, (c) following step (b) and prior to, simultaneous with, or following step (a), incubating the binding guide conjugates and half circle probes under conditions that promote ligation of half circle probes in pairs of half circle probes to each other, thereby producing one or more amplification target circles.
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138. A method comprising:
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(a) bringing into contact one or more analyte samples and one or more binding guide conjugates, wherein each binding guide conjugate comprises a specific binding molecule and a guide oligonucleotide, wherein each specific binding molecule interacts directly or indirectly with an analyte in the analyte sample, and incubating the analyte samples and the binding guide conjugates under conditions that promote interaction of the specific binding molecules and the analytes, (b) prior to, simultaneous with, or following step (a), bringing into contact the binding guide conjugates and one or more half circle probes, wherein each half circle probe comprises a single-stranded DNA molecule comprising two guide complement portions, wherein each guide complement portion is complementary to at least one of the guide oligonucleotides, and incubating the binding guide conjugates and the half circle probes under conditions that promote hybridization between the guide oligonucleotides and the half circle probes, (c) following steps (a) and (b), incubating the binding guide conjugates and half circle probes under conditions that promote ligation of half circle probes to each other, thereby producing one or more amplification target circles.
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Specification