Method and system for detection of nucleic acids

Abstract

In detecting a nucleic acid by using the nucleic acid amplification method, it is intended to shorten the time required for the measurement so as to systematically and efficiently examine a gene, etc. To achieve this object, use is made of a method of directly amplifying a nucleic acid by treating a collected biological sample wherein a means of inhibiting the degradation activity of the nucleic acid is introduced during or after the step of homogenizing the biological sample and thus the nucleic acid is directly amplified without isolating/purifying the nucleic acid component from the biological sample. In this method, the means of inhibiting the degradation activity of the nucleic acid is introduced at an acidic pH value (more specifically, from pH 2.5 to pH 5) with the use of a salt interacting with a substance inhibiting the nucleic acid amplification reaction and/or the nucleic acid component so as to directly amplify the nucleic acid without isolating/purifying the nucleic acid component.

1 - 30 . (canceled) 
     
     
         31 . A kit for direct nucleic acid amplification, which comprises:
 (A) a biological sample treatment solution having an acidic pH and comprising a surfactant, wherein the acidic pH is in the range of pH 2.5 to pH 5, and   (B) a primer for amplifying a target nucleotide sequence, the target nucleotide sequence being a marker selected from the group consisting of a cancer marker and a tumor associated marker.   
     
     
         32 . The kit of  claim 31 , wherein the biological sample treatment solution further comprises a nuclease inhibitor. 
     
     
         33 . The kit of  claim 32 , wherein the nuclease inhibitor is an RNase inhibitor. 
     
     
         34 . The kit of  claim 31 , wherein the biological sample treatment solution further comprises a salt which interacts with an inhibitor of nucleic acid amplification reaction. 
     
     
         35 . The kit of  claim 34 , wherein the salt is a chaotropic salt. 
     
     
         36 . The kit of  claim 31 , wherein said amplification is selected from the group consisting of LAMP, RT-LAMP, PCR and RT-PCR. 
     
     
         37 . The kit of  claim 31 , wherein the target nucleotide sequence is a nucleotide sequence selected from the group consisting of cytokeratin 18, cytokeratin 19, cytokeratin 20 and CEA. 
     
     
         38 . The kit of  claim 34 , wherein the salt is at least one member selected from the group consisting of NaCl, KCl, NaI, KI, TMACl (tetramethyl ammonium chloride), TEACl (tetraethyl ammonium chloride), KSCN, CsSCN and CsCl. 
     
     
         39 . The kit of  claim 31 , wherein the biological sample is a lymph node. 
     
     
         40 . The kit of  claim 31 , which further comprises a DNA polymerase. 
     
     
         41 . A combination of reagents, which comprises:
 (A) a biological sample treatment solution having an acidic pH and comprising a surfactant, wherein the acidic pH is in the range of pH 2.5 to pH 5, and   (B) a primer for amplifying a target nucleotide sequence, the target nucleotide sequence being a marker selected from the group consisting of a cancer marker and a tumor associated marker.   
     
     
         42 . The combination of  claim 41 , wherein the biological sample treatment solution further comprises a nuclease inhibitor. 
     
     
         43 . The combination of  claim 42 , wherein the nuclease inhibitor is an RNase inhibitor. 
     
     
         44 . The combination of  claim 41 , wherein the biological sample treatment solution further comprises a salt which interacts with an inhibitor of nucleic acid amplification reaction. 
     
     
         45 . The combination of  claim 44 , wherein the salt is a chaotropic salt. 
     
     
         46 . The combination of  claim 41 , wherein said amplifying a method selected from the group consisting of LAMP, RT-LAMP, PCR and RT-PCR. 
     
     
         47 . The combination of  claim 41 , wherein the target nucleotide sequence is a nucleotide sequence selected from the group consisting of cytokeratin 18, cytokeratin 19, cytokeratin 20 and CEA. 
     
     
         48 . The combination of  claim 44 , wherein the salt is at least one member selected from the group consisting of NaCl, KCl, NaI, KI, TMACl (tetramethyl ammonium chloride), TEACl (tetraethyl ammonium chloride), KSCN, CsSCN and CsCl. 
     
     
         49 . The combination of  claim 41 , wherein the biological sample is a lymph node. 
     
     
         50 . The combination of  claim 41 , which further comprises a DNA polymerase.