Isothermal Amplification of Nucleic Acids with Ladder-shape Melting Curve

A novel method termed isothermal amplification of nucleic acids with ladder-shape melting curve (LMCP) was developed in the study. In this method, one pair of primers or two pairs of nested primers and a thermostable DNA polymerase (large fragment) were employed to amplify the internal transcribed spacer (ITS) of Oryza sativa with ladder-shape melting curve. Our results demonstrated that the LMCP assay with nested primers was 50-fold more sensitive and one-hour faster than the LAMP assay with the same level of specificity. The LMCP method has the potential to be used for the prevention and control of the emerging epidemics.