DNA Amplifcation Based on Polymerase Chain Reaction for the Sensitive Detection of the Mycoplasmalike Organism Associated with Paulownia Witches'-Broom.

Abstract

DNA amplification by polymerase chain reaction (PCR) was employed to detect the mycoplasma-like organism (MLO) associated with paulownia witches'-broom (PaWB). A 1.2kb DNA fragment of PaWB MLO was amplified with primer pair R16F2R2. A minimum of only 16pg total nucleic acids from infected paulownia tissue culture maintained at 25-28°C was needed to detect PaWB MLO. An appropriate concentration of total nucleic acids for amplification used for detection of PaWB MLO from the tissue culture was demonstrated to be 240pg/μl to 6.5ng/μl. PaWB MLO DNA content in total nucleic acids from MLO-infected tissue culture was as high as 5 times that from MLO-infected paulownia plant grown at 23-25°C in the greenhouse. Results showed that the amplification by PCR is highly effective in detecting the low concentrations of PaWB MLO known to occur in irregular patterns in diseased paulownia plants.