Salmonella typhimurium X3306lux and applications of salmonella typhimurium X3306lux in living body imaging

A Salmonella typhimurium, in vivo imaging technology, applied in the field of microorganisms, can solve the problems of high background, low specificity of fluorescent labeling in vivo detection, difficult quantification, etc., achieve high specificity and signal-to-noise ratio, stable bioluminescence performance, The effect of a stable amount of light

Inactive Publication Date: 2012-10-31
SUZHOU UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0011] The present invention aims at the defects of low specificity, high background, and difficult quantification of fluorescent markers in vivo detection, and the shortcomings of conventional plasmid-based bioluminescence detection that require additional screening pressure and substrates, and are difficult to quantify, etc., and provides a stable luminescence Salmonella typhimurium X3306lux and its application in live imaging

Method used

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  • Salmonella typhimurium X3306lux and applications of salmonella typhimurium X3306lux in living body imaging
  • Salmonella typhimurium X3306lux and applications of salmonella typhimurium X3306lux in living body imaging
  • Salmonella typhimurium X3306lux and applications of salmonella typhimurium X3306lux in living body imaging

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Experimental program
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Embodiment 1

[0026] Specific construction and identification of strains:

[0027] 1. Material preparation

[0028] 1.1 Plasmids and strains

[0029] Plasmid pBEN276 was kindly donated by Prof. Pierre Germon, Laboratory of Animal Infectious Diseases and Public Health Pathogen Research, French Institute of Natural Resources. 【Kevin Howe, Attila Karsi, Pierre Germon, et.al. Development of stable reporter system cloning luxCDABE genes into chromosome of Salmonella enterica serotypes using Tn7 transposon [J]. BMC Microbiology 2010, 10:197】

[0030] Salmonella typhimurium ( Salmonella typhimurium ) X3306 was donated by Prof. ROY CURTISS III, School of Biological Sciences, Arizona State University, USA. [HIDENORI MATSUI, CHRISTOPHER M. BACOT, WENDY A. Virulence Plasmid-Borne spvB ​​and spvC Genes Can Replace the 90-Kilobase Plasmid in Conferring Virulence to Salmonella enterica Serovar Typhimurium in Subcutaneously Inoculated Mice[J]. Journal of bacteriology1, 5 (183): 4652–4658.]

[0031...

Embodiment 2

[0062] Salmonella typhimurium ( Salmonella typhimurium )Χ3306lux live imaging

[0063] 1. Bacterial culture

[0064] Select a single clone from the solid LB agar plate and inoculate it into LB liquid medium, and culture at 37°C, 200rpm for 14-16h to logarithmic growth phase. Collect bacteria by centrifugation at 4000rpm, 4°C, 10min, wash twice with normal saline (4000rpm, 4°C, 10min), resuspend with appropriate amount of normal saline, measure OD600 to calculate the bacterial concentration. Gradiently diluted with normal saline for use.

[0065] 2. Mice infection method (intraperitoneal injection or oral feeding infection)

[0066] 2.1 Intraperitoneal injection infection

[0067] Select 8-12 weeks BALB / C or C57BL mice, with 50μL concentration of 10 5 CFU / mL bacterial solution was injected intraperitoneally to infect mice.

[0068] 2.2. Oral feeding infection

[0069] After 8-12 weeks of BALB / C or C57BL mice were fasted for 6 hours, they were treated with 50 μL of 10%wt ...

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Abstract

The invention discloses a salmonella typhimurium X3306lux and applications of salmonella typhimurium X3306lux in living body imaging. The bacterial strain is preserved in depositary authority designated by the state intellectual property office, the preservation data is 13 March, 2012, the depositary authority is China general microbiological culture collection center, and the preservation number is CGMCC No.5895. The salmonella typhimurium X3306lux is a virulent strain carrying virulent plasmids pStSR100, can be taken as a standard strain, and is applied to the research of the pathogenic mechanism of salmonella typhimurium. X3306lux is the bacterial strain with stable bioluminescence property, which is constructed based on X3306. By the site-specific recombination, a housekeeping gene E.colifrr promoter and a Lux operon are inserted into chromosome of X3306 for stable expression (meanwhile, the insertion site does not damage the gene functions of a genome, and the bacteria can not suffer from the adverse influence). The Lux operon conducts constitutive expression on luciferase and a substrate under the regulation of the housekeeping gene E.colifrr promoter connected with the Lux operon at the upstream, the bacteria can light without additionally adding screening pressure and the substrate.

Description

[0001] technical field [0002] The invention belongs to the technical field of microorganisms, in particular to Salmonella typhimurium X3306lux and its application in live imaging. [0003] Background technique [0004] Salmonella typhimurium ( Salmonella typhimurium ) is an important zoonotic pathogen, which is mainly transmitted by water and food, and can cause food poisoning, gastroenteritis and other intestinal infectious diseases, and its infection incidence rate ranks first in Salmonella infection. X3306 is a virulent strain of Salmonella typhimurium carrying the virulence plasmid pStSR100, which can be used as a standard strain to study the pathogenic mechanism of Salmonella typhimurium. [0005] In recent years, in vivo optical imaging technology of living animals has been continuously developed in life science research due to its simple operation and intuitiveness. This imaging technology can directly observe the activities and responses of the marked genes and ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20A61K49/00C12R1/42
Inventor 黄瑞李嫄渊储元元叶颖
Owner SUZHOU UNIV
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