title: Rapid and Sensitive Isothermal Detection of Nucleic-acid Sequence by Multiple Cross Displacement Amplification.
abstract: We have devised a novel amplification strategy based on isothermal strand-displacement polymerization reaction, which was termed multiple cross displacement amplification (MCDA). The approach employed a set of ten specially designed primers spanning ten distinct regions of target sequence and was preceded at a constant temperature (61-65 °C). At the assay temperature, the double-stranded DNAs were at dynamic reaction environment of primer-template hybrid, thus the high concentration of primers annealed to the template strands without a denaturing step to initiate the synthesis. For the subsequent isothermal amplification step, a series of primer binding and extension events yielded several single-stranded DNAs and single-stranded single stem-loop DNA structures. Then, these DNA products enabled the strand-displacement reaction to enter into the exponential amplification. Three mainstream methods, including colorimetric indicators, agarose gel electrophoresis and real-time turbidity, were selected for monitoring the MCDA reaction. Moreover, the practical application of the MCDA assay was successfully evaluated by detecting the target pathogen nucleic acid in pork samples, which offered advantages on quick results, modest equipment requirements, easiness in operation, and high specificity and sensitivity. Here we expounded the basic MCDA mechanism and also provided details on an alternative (Single-MCDA assay, S-MCDA) to MCDA technique.
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Changbai Road 155, Changping, Beijing 102206, PR China; Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, PR China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Changbai Road 155, Changping, Beijing 102206, PR China; Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, PR China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Changbai Road 155, Changping, Beijing 102206, PR China; Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, PR China.
authors.forename: Ai-Jing
authors.initials: AJ
authors.lastname: Ma
authors.affiliation: Department of Microbiology, Guiyang Medical University, Guiyang, Guizhou 550004, PR China.
authors.forename: Dong-Xun
authors.initials: DX
authors.lastname: Li
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Changbai Road 155, Changping, Beijing 102206, PR China; Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, PR China.
authors.forename: Li-Juan
authors.initials: LJ
authors.lastname: Luo
authors.affiliation: Pathogenic Biology Institute, University of South China, Hengyang, Hunan 421000, PR China.
authors.forename: Dong-Xin
authors.initials: DX
authors.lastname: Liu
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Changbai Road 155, Changping, Beijing 102206, PR China; Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, PR China.
authors.forename: Dong
authors.initials: D
authors.lastname: Jin
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Changbai Road 155, Changping, Beijing 102206, PR China; Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, PR China.
authors.forename: Kai
authors.initials: K
authors.lastname: Liu
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Changbai Road 155, Changping, Beijing 102206, PR China; Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, PR China.
authors.forename: Chang-Yun
authors.initials: CY
authors.lastname: Ye
mesh_terms: D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D005516:Food Microbiology; D023061:Gene Order; D056426:Genetic Loci; D008089:Listeria monocytogenes / Q000235:genetics; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013696:Temperature
chemical_list: D017931:DNA Primers