An experimental study on HRM combined with multiplex PCR for the identification of single and mixed diatom species

Diatoms are research subjects in both ecological environments and forensic science. The identification of mixed diatom samples has always been a challenging issue. Multiplex polymerase chain reaction-high-resolution melting (PCR-HRM) provides a potential solution, but the effectiveness of this method in analyzing single and mixed diatom samples remains unclear. This study focuses on eight species of marine diatoms, identifying and designing corresponding primers to preliminarily explore the HRM technique combined with multiplex PCR for species identification of marine diatoms. By using two primer combinations, we achieved rapid identification of single diatoms. The experiments revealed that the HRM technique showed reduced identification capability for mixed diatom samples, whereas the multiplex PCR-HRM technique could enhance the identification accuracy for mixed diatom samples. By using a mixture of three primers, we identified four species of diatoms in mixed samples. This method can improve the identification capability of mixed diatom samples and provides a new approach for diatom species identification.