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Establishment of Multiplex PCR for Staphylococcu Saureus, Streptococcus, Salmonella Spp and Escherichia Coli

Author: YangLiHua
Tutor: XuXinGang；FanMingZhe
School: Northwest University of Science and Technology
Course: Veterinary
Keywords: Streptococcus Staphylococcus aureus Salmonella Escherichia coli Multiplex PCR
CLC: S855.1
Type: Master's thesis
Year: 2011
Downloads: 109
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Abstract


Streptococcus (Streptococcus), Staphylococcus aureus (Staphylococcus aureus), salmonella (Salmonella spp.) , And Escherichia coli ( Escherichia coli ) is widely distributed in nature, human public safety , animal health , food safety , and so has serious impact on the common pathogens causing human and livestock infected with the bacterial disease . Four kinds of bacteria that cause disease prevention and epidemiological investigations , as well as to understand the food safety status and the presence of the key issues are dependent on timely and accurate detection and identification of the pathogen . Therefore , this study for the NCBI has been included on the Streptococcus ef-tu gene , the complete genome sequence of Staphylococcus aureus nuc gene , Salmonella hut Gene and Escherichia coli 23SrRNA gene design and synthesis of four pairs of specific primers , by optimizing PCR amplification conditions multiplex PCR diagnostic method for the simultaneous detection of four bacterial infection . Such as annealing temperature and primer concentrations of the major factors that affect the multiplex PCR reaction appropriate adjustments and optimization to determine the optimal annealing temperature of 54 ° C for multiplex PCR the best primer concentration of 0.4 pmol / μL ; primer specificity analysis showed application of the method can be amplified from Streptococcus , Staphylococcus aureus , Salmonella and E. coli , as well as a mixture of four kinds of bacteria the four size was 197 bp , 278 bp, and 495 bp and 652 bp specific bands , but on the other control group test results were negative ; sensitivity analysis shows that the method detectable amount of genomic DNA of Four pathogens were Streptococcus 25.6 pg and Staphylococcus aureus 33.2 pg , Salmonella 35.7 pg, colorectal Aye Greek bacteria 52.1pg, this sensitivity is far higher than the the traditional bacteria were isolated and cultured detection method . Artificial mock-infected samples to detect multiple test results show that this method from mixed infection disease compound specific types of pathogenic bacteria detected . Multiplex PCR specificity , high sensitivity , good stability , simple and economical than traditional bacteriological methods can effectively detect Streptococcus , Staphylococcus aureus, Salmonella and Escherichia coli mixed infections . After further research and optimization can be used in of veterinary clinical diagnostics and food safety rapid detection .

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CLC: > Agricultural Sciences > Livestock, animal medicine,hunting,silkworm,bee > Animal Medicine ( Veterinary Medicine) > Veterinary Infectious Diseases > Bacterial Diseases
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