Design method of microbial multiplex PCR (Polymerase Chain Reaction) targeted amplification primer pool

The invention discloses a design method of a microorganism multiplex PCR (Polymerase Chain Reaction) targeted amplification primer pool, which aims at targeted PCR amplification of a large number of microorganisms, searches specific fragments of corresponding species at the in-species level, and designs a certain number of specific primer pairs. And designing a multiplex PCR targeted amplification primer pool based on a dimer scoring mechanism method for calculating a binary penalty score matrix and basic group mutual Gibbs free energy. The problems of high false positive and low speed of amplification of single targeted pathogenic microorganisms are solved. The multiple PCR primer pool can be efficiently designed, and the efficiency of simultaneous targeted amplification of a large number of microorganisms in a complex sample is improved. 本发明公开了一种微生物多重PCR靶向扩增引物池的设计方法，针对于大量微生物的靶向PCR扩增，寻找对应物种在种水平的特异性片段，并设计一定数量的特异性引物对，再基于二元罚分矩阵与碱基相互吉布斯自由能计算的二聚体评分机制方法，设计多重PCR靶向扩增引物池。解决了单重靶向病原微生物扩增假阳性较高，速度较慢的问题。可以高效的设计出多重PCR引物池，提