title: Quantitative phenotyping of X-disease resistance in chokecherry using real-time PCR.
abstract: A quantitative real-time SYBR Green PCR (qPCR) assay has been developed to detect and quantify X-disease phytoplasmas in chokecherry. An X-disease phytoplasma-specific and high sensitivity primer pair was designed based on the 16S rRNA gene sequence of X-disease phytoplasmas. This primer pair was specific to the 16SrIII group (X-disease) phytoplasmas. The qPCR method can quantify phytoplasmas from a DNA mix (a mix of both chokecherry and X-disease phytoplasma DNA) at as low as 0.001 ng, 10-fold lower than conventional PCR using the same primer pair. A significant correlation between the copy number of phytoplasmas and visual phenotypic rating scores of X-disease resistance in chokecherry plants was observed. Disease resistant chokecherries had a significantly lower titer of X-disease phytoplasmas than susceptible plants. This suggests that the qPCR assay provides a more objective tool to phenotype phytoplasma disease severity, particularly for early evaluation of host resistance; therefore, this method will facilitate quantitative phenotyping of disease resistance and has great potential in enhancing plant breeding.
authors.affiliation: Department of Plant Sciences, North Dakota State University, Fargo, ND 58108, USA.
authors.forename: Danqiong
authors.initials: D
authors.lastname: Huang
authors.affiliation: Department of Plant Pathology, North Dakota State University, Fargo, ND 58108, USA.
authors.forename: James A
authors.initials: JA
authors.lastname: Walla
authors.affiliation: Department of Plant Sciences, North Dakota State University, Fargo, ND 58108, USA. Electronic address: wenhao.dai@ndsu.edu.
authors.forename: Wenhao
authors.initials: W
authors.lastname: Dai
mesh_terms: D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D060467:Disease Resistance / Q000235:genetics*; D010641:Phenotype; D045582:Phytoplasma / Q000235:genetics*; D010935:Plant Diseases / Q000382:microbiology*; D027861:Prunus / Q000382:microbiology*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S
keywords: Candidatus Phytoplasma pruni; Quantitative phenotyping; Real-time PCR; X-disease resistance