Method for detecting multiple targets in same sample and method for quantifying concentration of multiple targets in same sample

A method for detecting multiple targets in the same sample, comprising:

• providing a sample, wherein the sample comprises a nucleic acid of at least one target;
• providing a reaction solution, wherein the reaction solution comprises:
• a primer pair set comprising a plurality of target primer pairs;
• and
• a fluorescent probe set comprising a plurality of target fluorescent probes and at least two fluorophores, and each of the at least two fluorophores being different, wherein each of the plurality of target fluorescent probes comprises a target oligonucleotide bound to at least one of the at least two fluorophores;
• wherein a number of the at least two fluorophores is m, a detectable number of the at least one target is 2m−1, a number of the plurality of target primer pairs is 2m−1, and a number of the plurality of target fluorescent probes is 2m−1;
• wherein each of the plurality of target primer pairs has sequence specificity with one of the at least one target, and the target oligonucleotide of each of the plurality of target fluorescent probes has sequence specificity with one of the at least one target;
• performing a reaction step, wherein the sample and the reaction solution are mixed and a real-time polymerase chain reaction is performed to obtain fluorescence signal values of the at least two fluorophores respectively;
• and
• performing a determining step, wherein a fluorescence signal value ratio of the at least two fluorophores is calculated to determine whether the at least one target is present in the sample.