A universal multiplex PCR strategy for 100-plex amplification using a hydrophobically patterned microarray

摘要: Both basic research and clinical medicine have urgent demands for highly efficient strategies to simultaneously identify many different DNA sequences within a single tube. Effective simultaneous amplification of multiple target is prerequisite any successful nucleic acid detection method. Multiplex PCR one the best choices this purpose. However, due intrinsic interference competition among primer pairs in same tube, rounds empirical optimization procedures are usually required establish multiplex reaction. To address challenge, we report here universal strategy that capable over 100-plex using specially designed microarray which hydrophilic microwells patterned on hydrophobic chip. On such an array, tagged with sequence physically separated individual otherwise chip, enabling unique reactions be proceeded during first step procedure. The products then isolated further amplified from sequences, producing sufficient amount material analysis by conventional gel electrophoresis or technology. This abbreviated as “MPH&HPM” “ultiplex CR ydrophobically ydrophilically atterned icroarray”. feasibility method demonstrated reaction eleven pneumonia-causing pathogens. Further, demonstrate power 116-plex only little prior optimization. effectiveness MPH&HPM samples illustrated deleted exons Duchenne Muscular Dystrophy (DMD) gene, results excellent agreement records. Because its generality, simplicity, flexibility, specificity capacity more than amplification, should broad applications both laboratory when required.