Method for multiplying nucleic acids

Abstract

The invention relates to a use of one or more nanoparticles, each of which is conjugated with at least one oligonucleotide, in order to multiply nucleic acids. One or more of the oligonucleotides has at least one primer sequence and an additional segment which extends from the end of the primer sequence proximal to the nanoparticle in the direction of the nanoparticle, and the additional segment has at least one abasic modification. The invention additionally relates to a method for multiplying nucleic acids in a sample, having a multiplication step for multiplying the nucleic acids and a test step for determining the concentration of the products of the multiplication reaction. The test step begins after the multiplication step ends, and in the test step, either at least one part of the sample is supplied with substances or no substances are supplied. The invention also relates to a method for multiplying nucleic acids, wherein nanoparticles transfer heat into their surroundings in a reaction volume upon being excited. The invention finally relates to a method for multiplying a nucleic acid using a polymerase chain reaction, in which a cycle consisting of a denaturing, annealing, and elongating step is repeatedly carried out.

1 . A nanoparticle for amplifying nucleic acids, wherein:
 the nanoparticle is conjugated to at least one oligonucleotide,   the oligonucleotide comprises at least one primer sequence and a further portion, the further portion extending from the nanoparticle-proximal end of the primer sequence in the direction of the nanoparticle, and   the further portion comprises at least one abasic modification.   
     
     
         2 . The nanoparticle of  claim 1 , wherein the abasic modification is arranged at the end, facing towards the primer sequence, of the further portion adjacently to the primer sequence. 
     
     
         3 . The nanoparticle of  claim 1 , wherein the abasic modification ( 7 ) is arranged 3′ sided with respect to the primer sequence. 
     
     
         4 . The nanoparticle of  claim 1 , wherein the abasic modification ( 7 ) is selected from the group including 1′,2′-dideoxyribose, triethylene glycol and hexaethylene glycol. 
     
     
         5 . The nanoparticle of  claim 1 , wherein the further portion comprises a plurality of abasic modifications ( 7 ). 
     
     
         6 . A method for amplifying nucleic acids in a sample comprising:
 an amplification step to amplify the nucleic acids; and
 a test step to determine the concentration of products of the amplification step, 
   wherein either:   no substances are added to the sample in the test step, or   the test step begins after the end of the amplification step, and substances are added at least to a part of the sample in the test step.   
     
     
         7 . (canceled) 
     
     
         8 . The method of  claim 6 , wherein a global temperature of the sample during the test step is different from a global temperature of the amplification step. 
     
     
         9 . The method of  claim 6 , wherein a global temperature of the sample during the test step is substantially equal to a global temperature of the amplification step. 
     
     
         10 . The method of  claim 6 , wherein, in the test step to determine concentration of products of the amplification step, at least one test probe is used, wherein each of the at least one test probes comprises a second nanoparticle. 
     
     
         11 . The method of  claim 10 , wherein the second nanoparticle of the test probe has a different size from that of the nanoparticle used in the amplification step to amplify the nucleic acids. 
     
     
         12 . A method for amplifying nucleic acids, wherein nanoparticles in a reaction volume transfer heat to their environment through excitation. 
     
     
         13 . The method of  claim 12 , wherein a duration of effect (t A ) is shorter than 10 s. 
     
     
         14 . The method of  claim 12 , wherein a duration of effect (t A ) is longer than 1 ps. 
     
     
         15 . The method of  claim 12 , wherein a heating time of the nanoparticles is shorter than 100 ms. 
     
     
         16 . The method of  claim 12 , wherein a cooling time is shorter than 100 ms. 
     
     
         17 . The method of  claim 12 , wherein a power density, with which the nanoparticles are excited, is more than 10 W/mm 2 . 
     
     
         18 . The method of  claim 12 , wherein a power density, with which the nanoparticles are excited, is less than 20,000 kW/mm 2 . 
     
     
         19 . The method of  claim 12 , wherein the concentration of the amplicon to be amplified in the method is greater than 10 −23  nM at the start of the method. 
     
     
         20 . The method of  claim 12 , wherein the concentration of the amplicon to be amplified in the method is less than 1 pM at the start of the method. 
     
     
         21 . The method of  claim 12 , wherein the number of amplicons to be amplified in the method is less than 500,000 at the start of the method. 
     
     
         22 . A method for amplifying a nucleic acid by means of a polymerase chain reaction, wherein a cycle of the polymerase chain reaction consisting of the steps denaturing, annealing and elongation is repeatedly passed through a number of passages, and wherein either:
 the number of passages of the cycle of the polymerase chain reaction is greater than 45; or   a cycle duration t c  is shorter than 40 seconds in at least one of the passages of the cycle of the polymerase chain reaction.   
     
     
         23 - 24 . (canceled)