The method and kit of multistage PCR detection NEISSERIA GONORRHOEAE

The method and kit of multistage PCR detection NEISSERIA GONORRHOEAE

  • CN 103,882,101 B
  • Filed: 10/17/2013
  • Issued: 11/30/2018
  • Est. Priority Date: 12/21/2012
  • Status: Active Grant
First Claim
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1. a kind of non-diagnostic and non-therapeutic type or type method for distinguishing of identifying the pathogen, which is characterized in that including step:

  • (a) in closed first order PCR reaction chamber or compartment, using contain or may the sample containing pathogen nucleic acid as mouldPlate, the first primer by the pathogen specific is to expanding, to obtain the first amplified production P1;

    (b) the first amplified production is transferred in the PCR compartment of the second level from first order PCR compartment, the mould as second level PCRPlate, and in closed second level PCR compartment, it is expanded by the second primer pair of the pathogen specific, to obtainObtain the second amplified production P2;

    (c) optionally, the amplified production Pi of previous step is transferred in i+1 grade PCR compartment from i-stage PCR compartment, asThe template of i+1 grade PCR, and in closed i+1 grade PCR compartment, pass through the i+1 primer pair of the pathogen specificIt is expanded, to obtain i+1 amplified production Pi+1, wherein i be >

    =2 positive integer;

    This step can carry out one or many;

    (d) to the second amplified production P2 or the i+1 amplified production P obtained in abovementioned stepsi+1, it is sequenced, fromAnd obtain the sequence of amplified production;

    With(e) the amplified production sequence measured is compared with pathogens standard sequence, thus identify pathogen type orType;

    Also, the pathogen is bacterium;

    And in step (a), it is placed with the solid particle that can carry pcr amplification product in first order PCR compartment, thus in PCRIn reaction process or later, the solid particle for carrying pcr amplification product is formed;

    And the solid particle is magnetic microsphere;

    AndIn step (b) and (c), jth is expanded by the solid particle for carrying pcr amplification product using magnet or electromagnetProduct PjIt is transferred to+1 grade of PCR compartment of jth from j-th stage PCR compartment, the positive integer that wherein j is >

    =1;

    Wherein, which carries out in multistage PCR reaction tube, and the reaction tube is by two or more closed PCR reaction chambersOr compartment (10), and the closed or closed interface channel (40) being equipped between at least two compartments formed, instituteState interface channel for allow carry pcr amplification product solid particle (50) from a compartment or upstream compartment be transferred to it is another everyRoom or downstream compartment;

    And the compartment has chamber lid, and for interconnected multiple compartments, when respective chamber lid all coversAfterwards, an enclosure space is just constituted, and the chamber lid is sealing cover;

    And start to multistage PCR to react to tie in multistage PCR reactionDuring beam, the entire multistage PCR reaction tube is in completely enclosed state.

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