title: Development of a rapid and high-throughput molecular method for detecting the F200Y mutant genotype in benzimidazole-resistant isolates of Fusarium asiaticum.
abstract: BACKGROUND
  The point mutation at codon 200 (TTC→TAC, F200Y) of the β2-tubulin gene confers resistance to benzimidazole fungicide in Fusarium asiaticum. These isolates with this mutation have been detected mainly by determining the minimum inhibitory concentration (MIC) of fungicides, which is always time consuming, tedious and inefficient.
  
  
  RESULTS
  A visual, rapid and efficient method with high specificity was developed, based on loop-mediated isothermal amplification (LAMP). Six sets of LAMP primers were designed, and one set was optimised specifically to distinguish the F200Y mutant genotype. With the optimal LAMP primers, concentrations of LAMP components were optimised. The optimal reaction conditions were 57-64 °C for 75 min. The feasibility of the LAMP assay for detection of the F200Y mutant genotype of F. asiaticum was demonstrated by assaying diseased wheat spikelets that were artificially inoculated in the field.
  
  
  CONCLUSIONS
  The new LAMP assay had good specificity, sensitivity, stability and repeatability. It will be useful for assessing the risk of F. asiaticum populations with carbendazim resistance developing in the field, and will also provide important reference data for integrated control of Fusarium head blight caused by F. asiaticum. © 2016 Society of Chemical Industry.
authors.affiliation: College of Plant Protection, State and Local Joint Engineering Research Centre of Green Pesticide Invention and Application, Nanjing Agricultural University, Nanjing, China.
authors.forename: Yabing
authors.initials: Y
authors.lastname: Duan
authors.affiliation: College of Plant Protection, State and Local Joint Engineering Research Centre of Green Pesticide Invention and Application, Nanjing Agricultural University, Nanjing, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Yang
authors.affiliation: College of Plant Protection, State and Local Joint Engineering Research Centre of Green Pesticide Invention and Application, Nanjing Agricultural University, Nanjing, China.
authors.forename: Tao
authors.initials: T
authors.lastname: Li
authors.affiliation: College of Plant Protection, State and Local Joint Engineering Research Centre of Green Pesticide Invention and Application, Nanjing Agricultural University, Nanjing, China.
authors.forename: Donglei
authors.initials: D
authors.lastname: Zhao
authors.affiliation: College of Plant Protection, State and Local Joint Engineering Research Centre of Green Pesticide Invention and Application, Nanjing Agricultural University, Nanjing, China.
authors.forename: Junhong
authors.initials: J
authors.lastname: Cao
authors.affiliation: College of Plant Protection, State and Local Joint Engineering Research Centre of Green Pesticide Invention and Application, Nanjing Agricultural University, Nanjing, China.
authors.forename: Yiyuan
authors.initials: Y
authors.lastname: Shi
authors.affiliation: College of Plant Protection, State and Local Joint Engineering Research Centre of Green Pesticide Invention and Application, Nanjing Agricultural University, Nanjing, China.
authors.forename: Jianxin
authors.initials: J
authors.lastname: Wang
authors.affiliation: College of Plant Protection, State and Local Joint Engineering Research Centre of Green Pesticide Invention and Application, Nanjing Agricultural University, Nanjing, China. mgzhou@njau.edu.cn.
authors.forename: Mingguo
authors.initials: M
authors.lastname: Zhou
mesh_terms: D001562:Benzimidazoles / Q000494:pharmacology*; D025141:Drug Resistance, Fungal; D005659:Fungicides, Industrial / Q000494:pharmacology*; D005670:Fusarium / Q000187:drug effects* / Q000235:genetics; D005838:Genotype; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D017354:Point Mutation
chemical_list: D001562:Benzimidazoles; D005659:Fungicides, Industrial; C031000:benzimidazole
keywords: Fusarium asiaticum; carbendazim; detection; loop-mediated isothermal amplification; resistance