RET fusion gene ARMS (amplification refractory mutation system) fluorescent quantitative PCR typing detection kit
A fusion gene and fluorescence quantitative technology, which is applied in the determination/inspection of microorganisms, DNA/RNA fragments, recombinant DNA technology, etc., can solve problems such as the inability to obtain amplification products, and achieve simplification of experiments, improvement of signal-to-noise ratio, and optimization steps. simple effect
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Embodiment 1
[0120] Example 1: Detection of RET fusion gene KR1 (KIF5B exon 15 / RET exon 12 fusion)
[0121] Use the fluorescent quantitative reaction solution A in the invented RET fusion gene ARMS fluorescent quantitative PCR typing detection kit for detection; then optimize the reaction system for ARMS-qPCR detection: the reaction system is 25μL, fluorescent quantitative reaction solution A (primers, probes) Mixture, concentration of 5uM) 1μL, sample template DNA 2μL (100-300ng / μL), 2*TaqmanuniversalPCRMasterMix (purchased from American Applied Biology) 15μL, ddH 2 O7μL.
[0122] PCR reaction conditions: 95°C pre-denaturation for 30 seconds; 95°C for 15 seconds, 62°C for 20 seconds, 20 cycles of amplification reaction; and 95°C for 15 seconds, 60°C for 34 seconds, and 40 cycles of amplification reaction.
[0123] At the same time, in the fluorescence quantitative test, while testing the sample, it is also necessary to set the sample reference product to determine the validity of the test, such ...
Embodiment 2
[0124] Example 2: Detection of RET fusion gene KR2 (KIF5B exon 16 / RET exon 12 fusion)
[0125] Use the fluorescent quantitative reaction solution A in the invented RET fusion gene ARMS fluorescent quantitative PCR typing detection kit for detection; then optimize the reaction system for ARMS-qPCR detection: the reaction system is 25μL, fluorescent quantitative reaction solution A (primers, probes) Mixture, concentration of 5uM) 1μL, sample template DNA 2μL (100-300ng / μL), 2*TaqmanuniversalPCRMasterMix (purchased from American Applied Biology) 15μL, ddH 2 O7μL.
[0126] PCR reaction conditions: 95°C pre-denaturation for 30 seconds; 95°C for 15 seconds, 62°C for 20 seconds, 20 cycles of amplification reaction; and 95°C for 15 seconds, 60°C for 34 seconds, and 40 cycles of amplification reaction.
[0127] At the same time, in the fluorescence quantitative test, while testing the sample, it is also necessary to set the sample reference product to determine the validity of the test, such ...
Embodiment 3
[0128] Example 3: Detection of RET fusion gene KR3 (KIF5B exon 23 / RET exon 12 fusion)
[0129] Use the fluorescent quantitative reaction solution A in the invented RET fusion gene ARMS fluorescent quantitative PCR typing detection kit for detection; then optimize the reaction system for ARMS-qPCR detection: the reaction system is 25μL, fluorescent quantitative reaction solution A (primers, probes) Mixture, concentration of 5uM) 1μL, sample template DNA 2μL (100-300ng / μL), 2*TaqmanuniversalPCRMasterMix (purchased from American Applied Biology) 15μL, ddH 2 O7μL.
[0130] PCR reaction conditions: 95°C pre-denaturation for 30 seconds; 95°C for 15 seconds, 62°C for 20 seconds, 20 cycles of amplification reaction; and 95°C for 15 seconds, 60°C for 34 seconds, and 40 cycles of amplification reaction.
[0131] At the same time, in the fluorescence quantitative test, while testing the sample, it is also necessary to set the sample reference product to determine the validity of the test, such ...
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