Serratia marcescens bacteriophage and medical application thereof
A technology of Serratia marcescens and phage, applied in the field of bioengineering, can solve the problem of less research on Serratia marcescens phage
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0020] Phage isolation and preparation
[0021] The isolation procedure of phage is described in detail below. The sewage sample in the present invention was collected from Huoshao Li in Changchun City, and the host bacterium Serratia marcescens SM2 was isolated from the First Hospital of Jilin University. Use collected sewage instead of double distilled water to prepare LB medium (100mL); add 1mL of overnight cultured host bacteria SM2 to the medium, and incubate at 37°C for 10-12h; take 1mL of the mixed culture, centrifuge at 12000×g for 5min, and use the supernatant The stock solution of phage was filtered through a 0.22 μm sterile filter and stored, and the obtained filtrate was used for plaque test to check whether the filtrate contained phage capable of lysing Serratia marcescens SM2.
[0022] Plaque test: Take Serratia marcescens SM2 cultured overnight at 37°C with shaking, and spread the bacterial solution evenly on the LB solid plate with a sterile cotton swab; after...
Embodiment 2
[0025] Phage amplification and purification
[0026] On the double-layer plate forming plaques, use a sterile pipette tip to pick up a large, round and translucent single plaque, and inoculate it in 5 ml LB liquid medium containing 200 μL of host bacterial solution Mix well, shake and culture at 37°C for 3-5 hours, then centrifuge at 12000×g, 4°C for 10 minutes, and take the supernatant; repeat the double-layer plate experiment, and repeat this operation 4-5 times until the cells of uniform size can be formed on the double-layer plate. of phage plaques.
[0027] Mix 5 mL of freshly cultivated host bacteria with 1 mL of phage lysate (according to the optimal MOI), add to 800 mL of LB liquid medium, and then add CaCl at a final concentration of 1.25 mM 2 , shake culture at 37°C for 6-8h, centrifuge at 12000×g for 10min at 4°C, and take the supernatant, which is the phage lysate.
[0028] PEG purification: Add RNase A and DNase I to the phage lysate to a final concentration of ...
Embodiment 3
[0035] Transmission electron microscope observation of phage vB_SamS-WG03
[0036] Take the phage purified by PEG-8000 in Example 2 for electron microscope observation. The specific operation steps are: add 10 μL of sample and drop it on the copper grid, wait for it to settle for 15 minutes, absorb the excess liquid with filter paper, and use 2% phosphotungstic acid ( PTA) was stained for 1-2min, and observed with a transmission electron microscope (Hitachi H-7650) after drying; the observation results were as follows figure 2 As shown, the head is an icosahedron, the diameter of the head is about 50±3nm, and the length of the tail is about 92±3nm. According to the "Classification of Viruses—The Eighth Report of the International Committee on Taxonomy of Viruses" published by the International Committee on Taxonomy of Viruses (ICTV) in 2005, vB_SamS-WG03 belongs to the family Siphoviridae.
PUM
| Property | Measurement | Unit |
|---|---|---|
| diameter | aaaaa | aaaaa |
Abstract
Description
Claims
Application Information
Login to View More