title: [HAND system-based four multiplex RT-PCR for simultaneous detection of four diarrhea viruses].
abstract: OBJECTIVE
  To establish a one-step four multiplex reverse transcription polymerase chain reaction (RT-PCR) method based on Homo-Tag Assisted Non-Dimer System (HAND) system for simultaneous detection of 4 diarrhea viruses of rotavirus, astrovirus, norovirus and sapovirus.
  
  
  METHODS
  Primers were designed according to the conserved genome sequence of the 4 viruses and the homologous tail sequences were added to the 5' end. The multiplex RT-PCR system was constructed by optimizing the PCR parameters such as the concentration of universal tag primer and genome-specific Homo-Tailed primers. The specificity, stability and sensitivity of the method were evaluated systematically.
  
  
  RESULTS
  The 4 multiplex RT-PCR methods based on HAND system was established successfully. Specificity analysis showed no cross reaction between the 4 diarrhea viruses. The sensitivity analysis showed detection limits for rotavirus, astrovirus, norovirus and sapovirus of 48, 1.92, 9.6 and 48 pg per reaction, respectively.
  
  
  CONCLUSIONS
  The established HAND system-based multiplex RT-PCR assay allows simple, rapid, specific, sensitive, and stable for detection of the 4 common diarrhea viruses at low costs and is suitable for application in general medical laboratories.
authors.affiliation: Department of Epidemiology, Southern Medical University, Guangzhou, China. gztengyy@163.com
authors.forename: Yongyong
authors.initials: Y
authors.lastname: Teng
authors.forename: Qiuhua
authors.initials: Q
authors.lastname: Mo
authors.forename: Qi
authors.initials: Q
authors.lastname: Wang
authors.forename: Minghui
authors.initials: M
authors.lastname: Tang
authors.forename: Dejian
authors.initials: D
authors.lastname: Zhao
authors.forename: Hua
authors.initials: H
authors.lastname: Tan
authors.forename: Chengning
authors.initials: C
authors.lastname: Tu
authors.forename: Ze
authors.initials: Z
authors.lastname: Yang
authors.forename: Qing
authors.initials: Q
authors.lastname: Chen
authors.forename: Hong
authors.initials: H
authors.lastname: Sun
mesh_terms: D019200:Astroviridae / Q000235:genetics / Q000302:isolation & purification*; D003967:Diarrhea / Q000821:virology*; D005243:Feces / Q000821:virology; D006801:Humans; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D029322:Norovirus / Q000235:genetics / Q000302:isolation & purification*; D012367:RNA, Viral / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D012401:Rotavirus / Q000235:genetics / Q000302:isolation & purification*; D029341:Sapovirus / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D012367:RNA, Viral