Construction and evaluation of reference standards for detection and quantification of Klebsiella pneumoniae using real-time PCR

作     者：Fei-Long Sun1,2,Min Jin3,Zhi-Gang Qiu3,Zhi-Qiang Shen3,Xin-Wei Wang3,Jun-Wen Li31.School of Life Science and Technology,Xi’an Jiaotong University,Xi’an 710049 2.School of Environmental and Chemical Engineering,Xi’an Polytechnic University,Xi’an 710048 3.Institute of Environment and Health,Tianjin 300050,China 

作者机构：School of Life Science and Technology Xi'an Jiaotong University Xi'an 710049 China School of Environmental and Chemical Engineering Xi'an Polytechnic University Xi'an 710048 China Institute of Environment and Health Tianjin 300050 China 

出 版 物：《Journal of Pharmaceutical Analysis》 (药物分析学报（英文版）)

年 卷 期：2010年第22卷第3期

页      面：183-187页

学科分类：100208[医学-临床检验诊断学] 1002[医学-临床医学] 10[医学] 

基　　金：supported by the National High Technology Research and Development Program of China(863Program No.2006AA06Z408) 

主　　题：cloning vector Klebsiella pneumoniae real-time PCR standard 

摘      要：Objective To construct reference standards for detection and quantification of Klebsiella pneumoniae(***)with SYBR Green I-based real-time PCR *** Primers were designed based on the published sequence of the phoE gene of *** standard was prepared by cell culture,PCR and T-A clone methods,and was identified by colony PCR and DNA *** The standard curve showed a very good linear negative regression between threshold cycle(Ct)and Log starting quantity of copy *** detection range was from 5.2 to 5.2×106 copies per reaction,and the detection limit was 6 copies per *** coefficients of variance(CVs)of three parallel experiments were in the range of 0.05%-0.91%.Conclusion The reference standards have high stability and *** can be used in the quantitative detection of ***.