Rapid and high-throughput identification of recombinant bacteria with mass spectrometry assay.

Di Xiao; Xiao Xia Tao; Peng Wang; Guo Dong Liu; Ya Nan Gong; Hui Fang Zhang; Hai Bin Wang; Jian Zhong Zhang

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Rapid and high-throughput identification of recombinant bacteria with mass spectrometry assay.

Author: Di XIAO ^{1
,

2} ; Xiao Xia TAO ¹ ; Peng WANG ³ ; Guo Dong LIU ⁴ ; Ya Nan GONG ¹ ; Hui Fang ZHANG ¹ ; Hai Bin WANG ⁵ ; Jian Zhong ZHANG ¹ ;
Author Information

1. State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China
2. Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou 310003, Zhejiang, China.
3. Provincial Key Laboratory for Plague Control and Prevention, Yunnan Provincial Institute for Endemic Diseases Control and Prevention, Dali 671000, Yunnan, China.
4. Beijing Municipal Center for Disease Control and Prevention, Beijing 102206, China.
5. Chaoyang District Center for Disease Control and Prevention, Beijing 100021, China.
Publication Type:Journal Article
Keywords: ClinProTools; MALDI-TOF MS; Rapid identification; Recombinant bacteria; Specific peaks
MeSH: Bacterial Proteins; analysis; Cloning, Molecular; Escherichia coli; Mass Spectrometry; Organisms, Genetically Modified; Peptide Mapping; Recombinant Proteins; analysis
From: Biomedical and Environmental Sciences 2014;27(4):250-258
CountryChina
Language:English
Abstract:
OBJECTIVETo construct a rapid and high-throughput assay for identifying recombinant bacteria based on mass spectrometry.
METHODSMatrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) techniques were used to identify 12 recombinant proteins (10 of Yersinia pestis, 1 of Campylobacter jejuni and 1 of Helicobacter pylori). A classification model for the various phase of recombinant bacteria was established, optimized and validated, using MALDI-TOF MS-ClinProTools system. The differences in the peptide mass spectra were analyzed by using Biotyper and FlexAnalysis softwares.
RESULTSModels of GA, SNN, and QC were established. After optimizing the parameters, the GA recognition model showed good classification capabilities: RC=100%, mean CVA=98.7% (the CVA was 96.4% in phase 1, 100% in phase 2, 98.4% in phase 3, and 100% in phase 4, respectively) and PPV=95%. This model can be used to classify the bacteria and their recombinant, which only requires 3.7×103 cells for analysis. The total time needed is only 10 min from protein extraction to reporting the result for one sample. Furthermore, this assay can automatically detect and test 96 samples concurrently. A total of 48 specific peaks (9, 16, 9, and 14 for the four stages, respectively) was found in the various phase of recombinant bacteria.
CONCLUSIONMALDI-TOF MS can be used as a fast, accurate, and high-throughput method to identify recombinant bacteria, which provide a new ideas not only for recombinant bacteria but also for the identification of mutant strains and bioterrorism pathogens.