Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Amino acid sequence and nucleotide sequence specifically combined with mycobacterium tuberculosis and application thereof

A technology of mycobacterium tuberculosis and nucleotide sequences, which is applied in the direction of medical preparations with non-active ingredients, non-active ingredients of polymer compounds, antibacterial drugs, etc., and can solve the problems of time-consuming, technically difficult, and false positives in culture methods To achieve the effects of improving diagnostic sensitivity and specificity, easy storage and transportation, and simplifying the preparation process

Active Publication Date: 2020-06-16
GENERAL HOSPITAL OF NINGXIA MEDICAL UNIV
View PDF4 Cites 1 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Early diagnosis is the first step in the treatment and termination of tuberculosis. However, traditional biological techniques such as microscopy and culture methods have shortcomings such as false positives, easy contamination, time-consuming, and inability to quantify, making it difficult to carry out in grassroots laboratories; With the continuous development of technology, technologies such as fully automatic real-time fluorescent nucleic acid amplification have been derived, but there are also defects such as high cost and technical difficulty, which cannot be widely used
In general, the existing laboratory diagnostic methods still have many limitations, and it is necessary to find a fast, accurate, economical and convenient diagnostic method

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Amino acid sequence and nucleotide sequence specifically combined with mycobacterium tuberculosis and application thereof
  • Amino acid sequence and nucleotide sequence specifically combined with mycobacterium tuberculosis and application thereof
  • Amino acid sequence and nucleotide sequence specifically combined with mycobacterium tuberculosis and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1: Amplification and purification of phage library

[0037]Inoculate a single colony of E.coli ER2738 in 5-10ml LB liquid medium, incubate at 37°C, 200rpm shaker until mid-logarithmic phase (OD600≈0.5); add 10ul phage (from NEB company), 37°C, 200rpm shaker Shake for 4-5 hours, centrifuge at 10,000 g for 10 min, and take the supernatant; centrifuge again at 10,000 g for 10 min, take 80% of the supernatant, add 1 / 6 volume of PEG8000 / NaCl, and stand overnight at 4°C. Take it out the next day, and the white precipitate is the phage. Centrifuge at 10,000g for 15 minutes, discard the supernatant, and gently suck out the residual solution after brief centrifugation; add 1ml TBS solution to dissolve the white precipitate.

Embodiment 2

[0038] Example 2: Panning and Identification of Heptapeptides Binding to M.tb

[0039] (1) Phage titer determination

[0040] Inoculate a single colony of E.coli ER2738 in 5-10ml LB liquid medium, incubate at 37°C, 250rpm shaker until mid-logarithmic phase (OD600≈0.5); heat and melt the top layer of agarose medium in a microwave oven, divide into 3mL / portion In the sterilized test tube, use one tube for each phage dilution, and store it at 45°C for later use; pre-warm the LB / IPTG / Xgal agar plate at 37°C, and take one plate for each phage dilution gradient; Doubling serial dilution (dilution range: amplified phage culture supernatant: 10 8 -10 11 ; Unamplified panning eluates: 10 1 -10 4 ); change a new tip for each dilution, and use a filter tip to avoid cross-contamination; when the E. Use one tube; add 10 μL of phages with different dilutions to each tube of E. coli liquid, shake and mix quickly, and incubate at room temperature for 1-5 minutes; add the phage-infected E...

Embodiment 3

[0047] Example 3: Identification of M.tb-specific phage

[0048] Randomly pick 20 well-separated single colonies on the titer determination plate after the third round of screening, culture and purify them separately, and use phage ELISA to detect the binding activity of each strain of phage to M.tb. The specific steps are as follows: ELISA plates were coated with M.tb and Blocking respectively, and each group was replicated three times, and blocked overnight at 4°C, washed 6 times with TBST, added 100 μL of purified phage, incubated at 37°C for 1 hour with shaking, TBST Wash 10 times to remove unbound phage, add 100 μL of HRP-labeled mouse anti-M13 monoclonal antibody, incubate with shaking at 37°C for 1 hour, wash 10 times with TBST, add TMB substrate for 5-10 minutes at room temperature in the dark, and stop the reaction with 2 mol / L sulfuric acid , Measure the OD value at a wavelength of 450nm, and take P / N ≥ 2.1 as positive.

[0049] The results of the P-ELISA detection ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses an amino acid sequence specifically combined with mycobacterium tuberculosis or a derivative of the amino acid sequence. The amino acid sequence or the derivative of the aminoacid sequence contains an amino acid sequence shown as SEQ ID NO: 1. The invention also discloses a nucleotide sequence for encoding the amino acid and application of the amino acid sequence. The amino acid sequence disclosed by the invention can be specifically combined with mycobacteria and is used for diagnosing tuberculosis, the diagnosis sensitivity and specificity are improved, and a new method is provided for targeted diagnosis of tuberculosis.

Description

technical field [0001] The invention belongs to the field of biotechnology, and particularly relates to an amino acid sequence, a nucleotide sequence and an application that specifically bind to Mycobacterium tuberculosis. Background technique [0002] Tuberculosis (TB), caused by Mycobacterium tuberculosis (M.tb) infection, is a worldwide infectious disease that seriously threatens public health. In terms of tuberculosis treatment, although most patients can recover through drug treatment, there are still a small amount of resident bacteria in the body after recovery. When the body is in a state of immunosuppression or low immunity, the resident bacteria will proliferate in large numbers, leading to the occurrence of tuberculosis again . With the emergence of multi-drug resistant strains of M.tb in recent years, the cross-infection of tuberculosis pathogens, the increase of population flow and the neglect of tuberculosis control, etc., the prevention and control of tubercu...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C07K7/06C12N15/11G01N33/569A61K47/42A61K47/64A61P31/06
CPCC07K7/06G01N33/5695A61K47/42A61K47/64A61P31/06
Inventor 薛頔魏萌萌林雪王玉炯
Owner GENERAL HOSPITAL OF NINGXIA MEDICAL UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com