US 11,859,243 B2
Differentiation of signals for target nucleic acid sequences
Jong Yoon Chun, Seoul (KR); and Young Jo Lee, Seoul (KR)
Assigned to SEEGENE, INC., Seoul (KR)
Filed by SEEGENE, INC., Seoul (KR)
Filed on Dec. 22, 2020, as Appl. No. 17/130,887.
Application 17/130,887 is a continuation of application No. 15/534,643, granted, now 10,876,155, previously published as PCT/KR2015/013461, filed on Dec. 9, 2015.
Claims priority of provisional application 62/154,319, filed on Apr. 29, 2015.
Prior Publication US 2021/0108260 A1, Apr. 15, 2021
This patent is subject to a terminal disclaimer.
Int. Cl. C12Q 1/6851 (2018.01); G16B 40/00 (2019.01); G16B 40/10 (2019.01)
CPC C12Q 1/6851 (2013.01) [G16B 40/00 (2019.02); G16B 40/10 (2019.02)] 26 Claims
 
1. A method for differentiating signals of interest for each of two target nucleic acid sequences comprising a first target nucleic acid sequence (T1) and a second target nucleic acid sequence (T2) in a sample, which are not differentiable by a single type of detector, comprising:
(a) incubating the sample with a first signal-generating means for detection of the first target nucleic acid sequence (T1) and a second signal-generating means for detection of the second target nucleic acid sequence (T2) and detecting signals at a first detection temperature (D1) and a second detection temperature (D2); wherein the signals of interest to be generated by the two signal-generating means are not differentiated for each target nucleic acid sequence by a single type of detector;
(b) providing the following two equations each of which comprises variables representing the signals of interest generated at each detection temperature for the two target nucleic acid sequences;
ST1D1+ST2D1=SD1  (I)
ST1D2+ST2D2=SD2  (II)
wherein (SD1) is a signal detected at the first detection temperature, (SD2) is a signal detected at the second detection temperature; (ST1D1) is a variable representing a signal of interest generated by the first signal-generating means at the first detection temperature, (ST2D1) is a variable representing a signal of interest generated by the second signal-generating means at the first detection temperature, (ST1D2) is a variable representing a signal of interest generated by the first signal-generating means at the second detection temperature, (ST2D2) is a variable representing a signal of interest generated by the second signal-generating means at the second detection temperature; and the total number of variables is four;
(c) providing two additional equations selected from the group consisting of the following equations:
f(ST1D1,ST1D2)=RVT1(D1D2)  (III),
f(ST2D1,ST2D2)=RVT2(D1D2)  (IV), and
ST1D2=0  (V)
wherein, RVT1(D1D2) is a reference value (RV) of the first target nucleic acid sequence (T1) representing a relationship of change in signals provided by the first signal-generating means at the first detection temperature and the second detection temperature, RVT2(D1D2) is a reference value (RV) of the second target nucleic acid sequence (T2) representing a relationship of change in signals provided by the second signal-generating means at the first detection temperature and the second detection temperature; f(ST1D1, ST1D2) represents a function of ST1D1 and ST1D2; f(ST2D1, ST2D2) represents a function of ST2D1 and ST2D2;
wherein ST1D2=0 in the equation (V) may be selected with a proviso that the first signal-generating means is prepared to generate no signal in the presence of the first target nucleic acid sequence at the second detection temperature; and
(d) obtaining solutions to at least one of the variables by the four equations provided in the steps (b) and (c) for differentiating at least one of the signals of interest to be assigned to at least one of the two target nucleic acid sequences.