Omega amplification

Abstract

The present disclosure provides compositions, methods and kits for Omega amplification technologies. In addition, the present disclosure provides compositions, methods and kits for universal FQ probe and for G-quadruplex detection methods for use in isothermal amplification technologies.

1 . A method of detecting a template nucleic acid in a sample using a strand displacement isothermal amplification reaction comprising
 (i) generating the reaction by combining the sample with (a) one or more amplification primers configured to generate amplicon nucleic acids from the template nucleic acids under suitable amplification conditions, and (b) a strand displacement amplification polymerase;   (ii) maintaining the reaction under the suitable amplification conditions; and   (iii) detecting whether amplification occurs or has occurred in step (ii) by monitoring during or after step (ii) interaction between (c) a specific detection probe that, under the suitable amplification conditions, hybridizes to the template nucleic acid, its compliment, the amplicon nucleic acid or its compliment, and (d) a universal detection probe.   
     
     
         2 . The method of  claim 1 , wherein the universal detection probe is a universal FQ probe. 
     
     
         3 . The method of  claim 1  or  claim 2 , wherein the universal detection probe does not anneal to the template nucleic acid or its complement under the suitable amplification conditions. 
     
     
         4 . The method of any one of  claims 1 - 3 , wherein the interaction between the specific detection probe and the universal detection probe is through hybridization during the amplification. 
     
     
         5 . The method of any one of  claims 1 - 3 , wherein the interaction between the specific detection probe and the universal detection probe is through hybridization between the complement of the specific detection probe and the universal detection probe. 
     
     
         6 . The method of any one of  claims 1 - 3 , wherein the interaction between the specific detection probe and the universal detection probe is through hybridization and polymerase extension during the amplification. 
     
     
         7 . The method of any one of  claims 1 - 3 , wherein the specific detection probe comprises an internal chemical moiety to stop polymerase extension. 
     
     
         8 . The method of any one of  claims 1 - 7 , wherein the universal detection probe comprises a first detection oligonucleotide strand and a second detection oligonucleotide strand. 
     
     
         9 . The method of  claim 8 , wherein (a) the first detection oligonucleotide strand comprises a quencher moiety and the second detection oligonucleotide strand comprises a fluorophore, or (b) the first detection oligonucleotide strand comprises a fluorophore and the second detection oligonucleotide strand comprises a quencher moiety, wherein the quencher moiety and the fluorophore are configured so that the quencher moiety quenches the fluorescence of the fluorophore when first detection oligonucleotide strand and a second detection oligonucleotide strand are annealed. 
     
     
         10 . The method of  claim 9 , wherein the ratio of the amount of the detection oligonucleotide strand comprising the fluorophore to the amount of the detection oligonucleotide strand comprising the quencher moiety is less than 1:1. 
     
     
         11 . The method of  claim 9  or  claim 10 , wherein the detecting step (ii) comprises measuring fluorescence emitted during the isothermal strand displacement amplification. 
     
     
         12 . The method of any one of  claims 8 - 11 , wherein the second detection oligonucleotide strand comprises an overhanging unmatched segment that is not complementary to the first detection oligonucleotide strand. 
     
     
         13 . The method of any one of  claims 8 - 12 , wherein the specific detection probe or its complement includes an invader that hybridizes to a portion of the overhanging unmatched segment and to a portion of the second detection oligonucleotide strand that is complementary to the first detection oligonucleotide strand during or after the amplification. 
     
     
         14 . The method of  claim 12 , further comprising an invader kicker probe includes mismatch near its 3′ end or at 3′ end when it hybridizes to the second detection oligonucleotide strand. 
     
     
         15 . The method of  claim 14 , further comprising an invader kicker replacement probe to replace the invader kicker probe once the invader kicker probe is extended along the second detection oligonucleotide strand. 
     
     
         16 . A method of detecting a template nucleic acid in a sample using a strand displacement isothermal amplification reaction comprising
 (i) generating the reaction by combining the sample with (a) one or more amplification primers configured to generate amplicon nucleic acids from the template nucleic acids under suitable amplification conditions, and (b) a strand displacement amplification polymerase;   (ii) maintaining the reaction under the suitable amplification conditions; and   (iii) detecting whether amplification occurs or has occurred in step (ii) by monitoring during or after step (ii) an aptamer probe; wherein the aptamer probe is part of a specific detection probe that, under suitable amplification conditions, hybridizes to the template nucleic acid, its compliment, an amplicon nucleic acid or its compliment.   
     
     
         17 . The method of  claim 16 , the aptamer probe is a G-quadruplex probe. 
     
     
         18 . The method of  claim 16  or  claim 17 , wherein the G-quadruplex probe generates a detectable signal selected from the group consisting of chromogenesis, fluorescence, luminescence, and chemiluminescence. 
     
     
         19 . The method of any one of  claims 1 - 18 , wherein the strand displacement amplification polymerase is selected from the group consisting of Bst DNA polymerase, Bca(exo-) DNA polymerase, Klenow fragment of DNA polymerase I, Vent DNA polymerase, Vent(Exo-) DNA polymerase (exonuclease activity-free Vent DNA polymerase), DeepVent DNA polymerase, DeepVent(Exo-) DNA polymerase (exonuclease activity-free DeepVent DNA polymerase), Φ29 phage DNA polymerase, MS-2 phage DNA polymerase, Z-Taq DNA polymerase (Takara Shuzo), and KOD DNA polymerase (TOYOBO). 
     
     
         20 . The method of any one of  claims 1 - 18 , wherein the strand displacement amplification polymerase is Bst DNA polymerase or Bca(exo-) DNA polymerase. 
     
     
         21 . The method of any one of  claims 1 - 20 , wherein one of the amplification primers is foldback primer. 
     
     
         22 . The method of any one of  claims 1 - 21 , wherein the strand displacement isothermal amplification reaction is LAMP, SMAP, GEAR, NEAR, or CPA. 
     
     
         23 . The method of any one of  claims 1 - 21 , wherein the isothermal amplification reaction is omega amplification and the pair of primers are foldback primers and at least one of the foldback primers is extruding primer. 
     
     
         24 . The method of  claim 23 , wherein the extruding sequence in the extruding primer comprises the specific detection probe sequences. 
     
     
         25 . The method of  claim 23  or  claim 24 , wherein the extruding sequence comprises internal modification to stop polymerase extension. 
     
     
         26 . The method of any one of  claims 1 - 25 , wherein the strand displacement isothermal amplification reaction comprises one or more than one kicker accelerator primers, or one or more than one stem accelerator primers, or one or more than one loop accelerator primers. 
     
     
         27 . The method of  claim 26 , wherein the kicker accelerator primer or loop accelerator primer or stem accelerator primer comprises folding sequences at its 5′ end to fold onto its 3′ end downstream sequences after 3′ end is extended by a polymerase. 
     
     
         28 . The method of any one of  claims 1 - 20 , wherein the strand displacement isothermal amplification reaction is RCA. 
     
     
         29 . The method of any one of  claims 1 - 20 , wherein the strand displacement amplification is nicking amplification and step (i) includes combining a nicking enzyme included in the reaction. 
     
     
         30 . The method of any one of  claims 1 - 29 , wherein the specific detection probe is an oligonucleotide that participates in the isothermal strand displacement amplification. 
     
     
         31 . An omega amplification primer set comprising a first foldback primer and a second foldback primer that allow isothermal amplification under suitable omega amplification conditions of a portion of a target nucleic acid sequence, wherein the first foldback primer comprises a first extruding sequence at its 5′ terminus or the second foldback primer comprises a second extruding sequence at its 5′ terminus. 
     
     
         32 . The omega amplification primer set of  claim 31 , wherein:
 (i) the target nucleic acid sequence has a first strand, wherein the first strand is complimentary to a complementary strand;   (ii) the first foldback primer includes from 5′ to 3′:
 (1-b) a sequence (F1c), wherein the sequence (F1c) hybridizes to a sequence (F1T) in the complimentary strand of the target nucleic acid sequence; and 
 (1-c) at the 3′ terminus, a sequence (F2), wherein the sequence (F2) hybridizes to a sequence (F2cT) in the first strand of the target nucleic acid sequence, 
 wherein the sequence (F1T) is 3′ of a sequence (F2T) in the complimentary strand; and the sequence (F2T) is complementary to the sequence (F2cT); 
   (iii) the second foldback primer includes from 5′ to 3′:
 (2-b) a second sequence comprising: a sequence (R1c), wherein the sequence (R1c) hybridizes to a sequence (R1T) in the first strand of the target nucleic acid sequence, 
 (2-c) at the 3′ terminus, a sequence (R2), wherein the sequence (R2) hybridizes to a sequence (R2cT) in the complimentary strand of the target nucleic acid sequence, 
 wherein the sequence (R1T) is 3′ of a sequence (R2T) in the first strand; and the sequence (R2T) is complementary to the sequence (R2cT); and 
   (iv) the primer set further comprises:
 (X) (1-a) a first extruding sequence at the 5′ terminus of the first foldback primer, wherein the first extruding sequence is at least 4 nucleotides and cannot hybridize to the first strand or the complimentary strand, and wherein the sequence (R1c) is at the 5′ terminus of the second foldback primer; 
 (Y) (2-a) a second extruding sequence at the 5′ terminus of the second foldback primer, wherein the second extruding sequence is at least 4 nucleotides and cannot hybridize to the first strand or the complimentary strand, and wherein the sequence (F1c) is at the 5′ terminus of the first foldback primer; or 
 (Z) (1-a) a first extruding sequence at the 5′ terminus of the first foldback primers, wherein the first extruding sequence is at least 4 nucleotides and cannot hybridize to the first strand or the complimentary strand, and (2-a) a second extruding sequence at the 5′ terminus of the second primer, wherein the second extruding sequence is at least 4 nucleotides and cannot hybridize to the first strand or the complimentary strand. 
   
     
     
         33 . The omega amplification primer set of  claim 32 , wherein a portion of the sequence (F1c) can hybridize to a portion of the sequence (R1c). 
     
     
         34 . The omega amplification primer set of  claim 32 , wherein the sequence (F1c) and the sequence (R1c) overlap by at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 16, 18, 20, 25, or 30 nucleotides. 
     
     
         35 . The omega amplification primer set of any one of  claims 31 - 34 , wherein the first extruding sequence or the second extruding sequence is at least 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 16, 18, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, 100, 150, or 200 nucleotides. 
     
     
         36 . The omega amplification primer set of any one of  claims 31 - 35 , wherein the first extruding sequence or the second extruding sequence is less than 500, 450, 400, 350, 300, 250, 200, 150, 100, 90, 80, 70, 60, 50, 45, 40, 35, 30, 25, or 20 nucleotides. 
     
     
         37 . The omega amplification primer set of any one of  claims 31 - 36 , wherein the first extruding sequence or the second extruding sequence is 3 to 100 nucleotides, 3 to 75 nucleotides, 3 to 50 nucleotides, or 4 to 30 nucleotides in length. 
     
     
         38 . The omega amplification primer set of any one of  claims 31 - 37 , wherein the first extruding sequence or the second extruding sequence comprises a G-quadruplex, an aptamer sequence, an RNA promoter sequence, a nicking sequence, or an FQ detection sequence. 
     
     
         39 . The omega amplification primer set of any one of  claims 31 - 38 , wherein the first extruding sequence or the second extruding sequence is G rich. 
     
     
         40 . The omega amplification primer set of any one of  claims 31 - 39 , wherein the omega amplification reaction comprises one or more than one kicker accelerator primers, or one or more than one stem accelerator primers, or one or more than one loop accelerator primers. 
     
     
         41 . The omega amplification primer set of any one of  claims 31 - 40 , wherein the kicker accelerator primer or loop accelerator primer or stem accelerator primer comprises folding sequences at its 5′ end to fold onto its 3′ end downstream sequences after 3′ end is extended by a polymerase. 
     
     
         42 . The omega amplification primer set of any one of  claims 31 - 41 , wherein the first extruding primer or the second extruding primer has hairpin structure at its 5′ terminus. 
     
     
         43 . The foldback primer amplification primer set of any one of  claims 31 - 42 , wherein foldback primer includes unnatural nucleotides. 
     
     
         44 . The foldback primer amplification primer set of any one of  claims 31 - 43 , wherein the folding hybridization sequence includes unnatural nucleotides. 
     
     
         45 . A method for determining whether a sample includes a template nucleic comprising
 (i) combining the sample with the set of omega amplification primers of any one of  claims 31 - 44 , a strand displacement amplification polymerase, and a detection probe; and   (ii) maintaining the combination under the suitable omega amplification conditions; and   (iii) determining whether the sample includes the template nucleic acid by monitoring whether the detection probe is involved in amplification during step (ii) or has been involved in amplification after step (ii).   
     
     
         46 . A method for assessing the amount of a template nucleic acid in a sample comprising
 (i) combining the sample with the set of omega amplification primers of any one of  claims 31 - 44 , a strand displacement amplification polymerase, and a detection probe; and   (ii) maintaining the combination under the suitable omega amplification conditions; and   (iii) quantifying the amount of the template nucleic acid by monitoring the detection probe during or after step (ii).   
     
     
         47 . The method of  claim 45  or  claim 46 , wherein the monitoring is performed during step (ii). 
     
     
         48 . The method of any one of  claims 45 - 47 , wherein the monitoring is based on a chromogenic reaction, a turbidity reaction, a chemiluminescent reaction, or a fluorescent reaction. 
     
     
         49 . The method of any one of  claims 45 - 47 , wherein the monitoring is monitoring fluorescent signal change from the detection probe. 
     
     
         50 . The method of any one of  claims 45 - 49 , wherein the detection probe has a universal FQ primer complement attached at its 5′ end. 
     
     
         51 . The method of any one of  claims 44 - 49 , wherein the detection probe is a specific detection probe and the monitoring is based on interaction between the specific detection probe or its complement and a universal FQ probe during amplification or after amplification. 
     
     
         52 . The method of  claim 51 , wherein the universal FQ probe comprises a first FQ oligonucleotide strand and a second FQ oligonucleotide strand. 
     
     
         53 . The method of  claim 52 , wherein the first FQ oligonucleotide strand and the second FQ oligonucleotide strand do not hybridize to the template strand under the suitable omega amplification conditions. 
     
     
         54 . The method of  claim 52  or  claim 53 , wherein (a) the first FQ oligonucleotide strand comprises a quencher moiety and the second FQ oligonucleotide strand comprises a fluorophore, or (b) the first FQ oligonucleotide strand comprises a fluorophore and the second FQ oligonucleotide strand comprises a quencher moiety, wherein the quencher moiety and the fluorophore are configured so that the quencher moiety quenches the fluorescence of the fluorophore when first FQ oligonucleotide strand and a second FQ oligonucleotide strand are annealed and the detecting comprises measuring fluorescence emitted during the isothermal strand displacement amplification. 
     
     
         55 . The method of  claim 54 , wherein the ratio of the amount of the FQ oligonucleotide strand comprising the fluorophore to the amount of the FQ oligonucleotide strand comprising the quencher moiety is less than 1:1 
     
     
         56 . The method of any one of  claims 52 - 55 , wherein the second FQ oligonucleotide strand comprises an overhanging unmatched segment that is not complementary to the first FQ oligonucleotide strand. 
     
     
         57 . The method of  claim 56 , wherein the specific detection probe or its complement includes an invader that hybridizes to a portion of the overhanging unmatched segment and to a portion of the second detection oligonucleotide strand that is complementary to the first detection oligonucleotide strand during or after the amplification. 
     
     
         58 . The method of  claim 57 , further comprising an invader kicker probe includes mismatch near its 3′ end or at 3′ end when it hybridizes to the second detection oligonucleotide strand. 
     
     
         59 . The method of  claim 58 , further comprising an invader kicker replacement probe to replace the invader kicker probe once the invader kicker probe is extended along the second detection oligonucleotide strand. 
     
     
         60 . The method of any one of  claims 45 - 59 , wherein the detection probe or universal detection probe includes a G-quadruplex probe or an aptamer probe. 
     
     
         61 . The method of any one of  claim 45 - 60 , wherein the first extruding sequence or the second extruding sequence comprises the detection probe. 
     
     
         62 . The method of any of  claims 45 - 61 , wherein the template nucleic acid is a human papilloma virus (HPV). 
     
     
         63 . The method of  claim 62 , wherein the HPV is HPV6, HPV11, HPV16, HPV18, HPV35, or HPV73. 
     
     
         64 . The method of  claim 62 , wherein the set of omega amplification primers are 18FIP (SEQ ID NO:1) and ex18BIP (SEQ ID NO:4), ex18FIP (SEQ ID NO:2) and 18BIP (SEQ ID NO:3), or ex18FIP (SEQ ID NO:2) and ex18BIP (SEQ ID NO:4), optionally including a kicker acceleration primer 18KF (SEQ ID NO:9) and/or 18 KB (SEQ ID NO:10), optionally including a loop acceleration primer 18LF (SEQ ID NO:5) and/or 18LB (SEQ ID NO:6), and optionally including an FQ probe comprising FAM-18LB (SEQ ID NO:7) and Q-oligo (SEQ ID NO:8). 
     
     
         65 . The method of  claim 62 , wherein the set of omega amplification primers are 73ovlp-exFIP (SEQ ID NO: 15) and 73-BIP (SEQ ID NO:18), 7350ovlp-exFIP (SEQ ID NO:16) and 73-BIP (SEQ ID NO:18), or 73-exFIP (SEQ ID NO:17) and 73-BIP (SEQ ID NO:18), optionally including a kicker acceleration primer 73-KF (SEQ ID NO:24) and/or 73-KB (SEQ ID NO:25), optionally including a loop acceleration primer 73ovlp-LF (SEQ ID NO:19), 7350ovlp-LF (SEQ ID NO:20), 73-LF (SEQ ID NO:21), and/or 73-LB (SEQ ID NO:22), and optionally including an FQ probe comprising Fam-73-LB (SEQ ID NO:23) and Q-oligo (SEQ ID NO:8). 
     
     
         66 . The method of  claim 62 , wherein the set of omega amplification primers are HPV6G-FIP (SEQ ID NO:27) and HPV6G BIP-22 nt (SEQ ID NO:29), optionally including a kicker acceleration primer HPV6G-KF (SEQ ID NO:33) and/or HPV6G-KB (SEQ ID NO:34), optionally including a loop acceleration primer 73ovlp-LF (SEQ ID NO:19), 7350ovlp-LF (SEQ ID NO:20), 73-LF (SEQ ID NO:21), and/or 73-LB (SEQ ID NO:22), and optionally including an FQ probe comprising Fam-73-LB (SEQ ID NO:23) and Q-oligo (SEQ ID NO:8). 
     
     
         67 . The method of  claim 62 , wherein the set of omega amplification primers are 35-exFIP (SEQ ID NO: 45) and 35-BIP (SEQ ID NO: 37), optionally including a kicker acceleration primer 35-KF (SEQ ID NO: 42) and/or 35-KB (SEQ ID NO: 43), optionally including a loop acceleration primer 35-LF (SEQ ID NO: 38), 35-FBLF (SEQ ID NO: 39), 35-LB (SEQ ID NO: 40), and/or 35-FBLB (SEQ ID NO: 41), and optionally including an FQ probe comprising 35-LF-FAM (SEQ ID NO: 44)) and Q-oligo (SEQ ID NO:8). 
     
     
         68 . A method of generating amplicon nucleic acids from a template nucleic acid in a sample using an omega amplification reaction comprising
 (i) combining the sample with the set of omega amplification primers of any one of  claims 31 - 44 , and a strand displacement amplification polymerase; and   (ii) generating amplicon nucleic acids by maintaining the combination under suitable omega amplification conditions.   
     
     
         69 . A method using the set of primers of any one of  claims 32 - 44  to make an amplicon nucleic acid from the target nucleic acid molecule, wherein the amplicon nucleic acid is capable of forming a first stem and loop at a first end, is capable of forming either a second stem and loop or a foldback loop at a second end, and has (i) the first extruding sequence located at the terminus of the first end, and/or (ii) the second extruding sequence located at the terminus of the second end, the method comprising:
 (a) combining a sample with the target nucleic acid molecule with the set of primers of any one of  claims 32 - 44 ; 
 (b) annealing the sequence (F2) of the first primer to the sequence (F2cT) in the first strand of the target nucleic acid molecule; 
 (c) extending the first primer from its 3′ end, using a suitable polymerase, to form a first single-stranded nucleic acid molecule comprising the first primer at the 5′ end and the sequence (R2cT); 
 (d) displacing the first single-stranded nucleic acid molecule from the target nucleic acid sequence; 
 (e) annealing the sequence (R2) of the second primer to the sequence (R2cT) in the first single-stranded nucleic acid molecule; and 
 (f) making the replicated portion of the target nucleic acid molecule by extending the second primer from its 3′ end, using a suitable polymerase, to form a second single-stranded nucleic acid molecule comprising the second primer at the 5′ end and a sequence complimentary to the first primer; 
 
       wherein the displacing step (d) is carried out by:
 (i) annealing the sequence (F2) of an additional first primer to the sequence (F2cT) in the first strand of the target nucleic acid molecule and extending the additional first primer from its 3′ end, using a suitable polymerase, to displace the first single-stranded nucleic acid molecule; 
 (ii) steps (d) and (e); or
 (iii) (1) providing a first kicker primer comprising, at its 3′ terminus, a sequence (F3), wherein the sequence (F3) hybridizes to a sequence (F3cT) and the sequence (F3cT) is 5′ of the sequence (F2cT) in the first strand of the target nucleic acid sequence;
 (2) annealing the sequence (F3) in the first kicker primer to the sequence (F3cT) in the first strand of the target nucleic acid molecule; and 
 (3) extending the first kicker primer from its 3′ end, using a suitable polymerase, to displace the first single-stranded nucleic acid molecule. 
 
 
 
     
     
         70 . The method of any one of  claims 45 - 69 , wherein the reaction is at least 20% as fast, at least 30% as fast, at least 40% as fast, at least 50% as fast, at least 60% as fast, at least 70% as fast, at least 80% as fast, or even at least 100% as fast as the same reaction where the first extruding primer does not comprises the first extruding sequence at its 5′ terminus and/or the second extruding primer does not comprise a second extruding sequence at its 5′ terminus. 
     
     
         71 . The method of any one of  claims 45 - 70 , wherein the first extruding sequence or the second extruding sequence is at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 16, 18, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, 100, 150, or 200 nucleotides. 
     
     
         72 . The method of any one of  claims 45 - 71 , wherein the first extruding sequence or the second extruding sequence is 1 to 100 nucleotides, 2 to 75 nucleotides, 3 to 50 nucleotides, or 4 to 30 nucleotides in length. 
     
     
         73 . The method of any one of  claims 45 - 72 , wherein the strand displacement amplification polymerase is selected from the group consisting of Bst DNA polymerase, Bca(exo-) DNA polymerase, Klenow fragment of DNA polymerase I, Vent DNA polymerase, Vent(Exo-) DNA polymerase (exonuclease activity-free Vent DNA polymerase), DeepVent DNA polymerase, DeepVent(Exo-) DNA polymerase (exonuclease activity-free DeepVent DNA polymerase), Φ29 phage DNA polymerase, MS-2 phage DNA polymerase, Z-Taq DNA polymerase (Takara Shuzo), and KOD DNA polymerase (TOYOBO). 
     
     
         74 . The method of any one of  claims 45 - 72 , wherein the strand displacement amplification polymerase is Bst DNA polymerase or Bca(exo-) DNA polymerase. 
     
     
         75 . The method of any one of  claims 45 - 74 , wherein the sample is selected from a specimen, a culture, a patient sample, a subject sample, a biological sample, and an environmental sample. 
     
     
         76 . The method of  claim 75 , wherein the patient sample or the subject sample is from blood, saliva, cerebral spinal fluid, pleural fluid, milk, lymph, sputum, semen, stool, swabs, Broncho Alveolar Lavage Fluid, tissue samples, or urine. 
     
     
         77 . The method of any one of  claims 45 - 76 , wherein the combining step further comprises combining with a reaction accelerator selected from the group consisting of one or more acceleration primers, an RNA polymerase promoter, a nicking sequence, and combinations thereof. 
     
     
         78 . The method of  claim 77 , wherein the reaction accelerator comprises the one or more acceleration primers and the acceleration primers are selected from the group consisting of kicker acceleration primers, loop acceleration primers, and stem acceleration primers. 
     
     
         79 . The method of  claim 77  or  claim 78 , wherein the reaction accelerator comprises the RNA polymerase promoter and the RNA polymerase promoter is included in the first extruding primer, the second extruding primer, the kicker acceleration primer, the loop acceleration primer, or the stem acceleration primer. 
     
     
         80 . The method of  claim 77 , wherein the RNA polymerase promoter is a T7 RNA polymerase promoter. 
     
     
         81 . The method of  claim 77 , wherein the reaction accelerator comprises the nicking sequence and the nicking sequence is included in the first extruding primer, the second extruding primer, the kicker acceleration primer, the loop acceleration primer, or the stem acceleration primer. 
     
     
         82 . A kit comprising the set of primers of any of  claim 31 - 44 . 
     
     
         83 . The kit of  claim 82 , further comprising a strand displacement amplification polymerase. 
     
     
         84 . The kit of  claim 83 , wherein the strand displacement amplification polymerase is selected from the group consisting of Bst DNA polymerase, Bca(exo-) DNA polymerase, Klenow fragment of DNA polymerase I, Vent DNA polymerase, Vent(Exo-) DNA polymerase (exonuclease activity-free Vent DNA polymerase), DeepVent DNA polymerase, DeepVent(Exo-) DNA polymerase (exonuclease activity-free DeepVent DNA polymerase), Φ29 phage DNA polymerase, MS-2 phage DNA polymerase, Z-Taq DNA polymerase (Takara Shuzo), and KOD DNA polymerase (TOYOBO). 
     
     
         85 . The kit of  claim 83 , wherein the strand displacement amplification polymerase is Bst DNA polymerase or Bca(exo-) DNA polymerase. 
     
     
         86 . The kit of any one of  claims 82 - 85 , further comprising a kicker acceleration primer, a loop acceleration primer, and/or a stem acceleration primer. 
     
     
         87 . The kit of any one of  claims 82 - 86 , further comprising a detection probe. 
     
     
         88 . The kit of  claim 87 , further comprising a universal detection probe that interacts with the detection probe during isothermal amplification. 
     
     
         89 . The kit of any one of  claims 82 - 88 , further comprising a thermostable luciferase, luciferin and an enzyme that converts inorganic pyrophosphate to ATP. 
     
     
         90 . An amplicon nucleic acid derived from a target nucleic acid sequence comprising from 5′ to 3′:
 (2) a second sequence comprising a sequence (R1c); 
 (3) a sequence (R2), wherein the sequence (R2) hybridizes to a sequence (R2cT) in a complimentary strand of the target nucleic acid sequence; 
 (4) a sequence (R1T), wherein the sequence (R1T) hybridizes to the sequence (R1c); 
 (5) a sequence (F1cT); 
 (6) a sequence (F2c), wherein the sequence (F2c) hybridizes to a sequence (F2T) in the complimentary strand of the target nucleic acid sequence; and 
 (7) a sequence (F1), wherein the sequence (F1) hybridizes to (F1cT) wherein the nucleic acid further comprises:
 (X) (8) a first extruding sequence at the 3′ terminus, wherein the first extruding sequence is at least 4 nucleotides and cannot hybridize to the template nucleic acid or its compliment, and wherein the sequence (R1c) is at the 5′ terminus; 
 (Y) (1) a second extruding sequence at the 5′ terminus, wherein the second extruding sequence is at least 4 nucleotides and cannot hybridize to the template nucleic acid or its compliment, and wherein the sequence (F1) is at the 3′ terminus; or 
 (Z) (8) a first extruding sequence at the 3′ terminus, wherein the first extruding sequence is at least 4 nucleotides and cannot hybridize to the template nucleic acid or its compliment, and (1) a second extruding sequence at the 5′ terminus, wherein the second extruding sequence is at least 4 nucleotides and cannot hybridize to the template nucleic acid or its compliment.