US 12,168,804 B2
Listeria-monocytogenes detection method
Mika Tsuna, Atsugi (JP); Yoshitaka Harada, Atsugi (JP); Yasuhiro Seto, Atsugi (JP); and Kazuto Takasaki, Atsugi (JP)
Assigned to NIPPN CORPORATION, Tokyo (JP); and FASMAC CO., LTD., Atsugi (JP)
Appl. No. 16/632,051
Filed by NIPPN CORPORATION, Tokyo (JP); and FASMAC CO., LTD., Atsugi (JP)
PCT Filed Dec. 18, 2017, PCT No. PCT/JP2017/045280
§ 371(c)(1), (2) Date Aug. 20, 2020,
PCT Pub. No. WO2019/016976, PCT Pub. Date Jan. 24, 2019.
Claims priority of application No. 2017-141201 (JP), filed on Jul. 20, 2017.
Prior Publication US 2021/0024979 A1, Jan. 28, 2021
Int. Cl. C12Q 1/689 (2018.01); C12Q 1/6853 (2018.01); C12Q 1/686 (2018.01); C12Q 1/6869 (2018.01); G01N 33/52 (2006.01)
CPC C12Q 1/689 (2013.01) [C12Q 1/6853 (2013.01); C12Q 1/686 (2013.01); C12Q 1/6869 (2013.01); G01N 33/52 (2013.01); C12Q 2600/156 (2013.01); G01N 2458/00 (2013.01); G01N 2469/00 (2013.01)] 9 Claims
 
1. A primer set for detection of Listeria monocytogenes, comprising any of the following primer sets for amplification of a partial region of lmo0084 gene or lmo2736 gene of Listeria monocytogenes:
(A-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:26 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:30 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(A-2) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:26 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:31 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(A-3) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:27 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:30 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(A-4) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:27 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:31 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(A-5) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:28 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in the base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:30 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(A-6) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:28 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:31 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(A-7) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:29 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:30 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(A-8) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:29 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:31 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(B-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:58 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:59 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(C-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:32 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:37 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(D-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:33 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:38 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(E-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:34 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in the base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:38 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(F-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:34 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:39 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(G-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:34 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:40 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(H-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:35 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in the base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:39 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(I-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:35 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:40 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(I-2) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:60 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:61 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(I-3) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:62 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:61 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(J-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:35 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:41 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence;
(K-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:36 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:39 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in the base sequence;
(L-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:36 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:40 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence; and
(M-1) a set of a forward primer containing in its 3′-side the base sequence of SEQ ID NO:63 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence, and a reverse primer containing in its 3′-side the base sequence of SEQ ID NO:64 or a sequence which is the same as said base sequence except that not more than 4 bases are substituted at a genetic polymorphism site(s) in said base sequence wherein the primer set further comprises a probe containing an oligonucleotide portion, wherein the ends of the oligonucleotide portion are modified with a fluorescent substance and a quencher substance, respectively.