Genomic DNA extraction method suitable for bacteria and/or fungi

An extraction method and genome technology, applied in the biological field, can solve the problems of prolonged detection time, inaccurate test results, low sensitivity, etc., and achieve the effects of reducing the possibility of errors, shortening the required time, and having a wide range of applications.

Inactive Publication Date: 2020-02-18
BEIJING BEIER BIOENG
View PDF8 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the need for in vitro culture, the required detection time is greatly prolonged; and the interference of inhibitory substances (such as antibacterial drugs) in clinical specimens often leads to the failure of in vitro culture of bacteria, which makes the test results inaccurate or low sensitivity, so that bacteria or Undetectable levels of fungi

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
  • Genomic DNA extraction method suitable for bacteria and/or fungi
  • Genomic DNA extraction method suitable for bacteria and/or fungi
  • Genomic DNA extraction method suitable for bacteria and/or fungi

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] 1. Process clinical specimens to obtain bacterial and / or fungal pellet concentrates

[0046] By processing different clinical specimens, bacteria and / or fungi can be effectively separated from somatic cells and body fluid components, so that bacteria can be concentrated to increase the unit effective concentration of bacteria and / or fungi. The processing methods for clinical specimens with different characteristics are given below:

[0047] For the whole blood sample, since the sample contains a large number of red blood cells, the heme released after the lysed red blood cells has a certain degree of inhibition on the subsequent DNA analysis, so for the whole blood sample, the operation of the present invention is: take 500 μl of anticoagulated blood, add 450 μl of red blood cells to lyse Solution (the composition of erythrocyte lysate is: 0.15M NH 4 Cl, 10mM KHCO 3 , 0.1mM EDTA, water, pH7.2-8.0) after inverting and mixing, let it stand for 5 minutes to lyse red bloo...

Embodiment 2

[0065] Example 2: Extraction and detection of Staphylococcus aureus genomic DNA in blood samples

[0066] Staphylococcus aureus belongs to the genus Staphylococcus, is a representative of Gram-positive bacteria, and is a common food-borne pathogenic bacteria.

[0067] A copy was tested positive for Staphylococcus aureus by the automatic biological mass spectrometry detection system Microflex LT / SH, and the bacterial content was determined to be 1.0×10 by DENSICHECK turbidimeter. 8 CFU / ml bacterial solution, use normal EDTA anticoagulated whole blood to dilute the bacterial solution to contain 1.0×10 6 CFU / ml (No. Sau106), 1.0×10 5 CFU / ml (code Sau105), 1.0×10 4 CFU / ml (No. Sau 104), 1.0×10 3 CFU / ml (code Sau103), 1.0×102CFU / ml (code Sau 102), 1.0×10 1 CFU / ml (No. Sau 101) blood samples of Staphylococcus aureus were used as experimental materials for the extraction of bacterial genomic DNA from blood as simulated bacterial infection samples.

[0068] Take more than 500ul s...

Embodiment 3

[0079] Example 3: Extraction and detection of Neisseria meningitidis genomic DNA in cerebrospinal fluid samples

[0080] Neisseria meningitidis (N. meningitidis), referred to as meningococcus (meningococcus), is the pathogenic bacteria of epidemic cerebrospinal meningitis (referred to as meningitis). Neisseria meningitidis is a Gram-negative diplococcus with a diameter of about 0.8 μm.

[0081] A portion was tested positive for Neisseria meningitidis by the automatic biological mass spectrometry detection system Microflex LT / SH, and the bacterial content measured by the DENSICHECK turbidimeter was 3.0×10 8 CFU / ml bacterial solution, respectively use the non-bacterial infected cerebrospinal fluid to dilute the bacterial solution to contain 5.0×10 5 CFU / ml (No. Nm105), 1.0×10 4 CFU / ml (No. Nm104), 1.0×10 3 CFU / ml (No. Nm 103), 1.0×10 2 CFU / ml (No. Nm 102) Neisseria meningitidis cerebrospinal fluid sample, as a simulated bacterial infection sample for genomic DNA extraction e...

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

Embodiments of the present invention relate to a genomic DNA extraction method suitable for bacteria and / or fungi. The method comprises the following steps: removing a liquid part in a clinical specimen to obtain a concentrate of fungi and / or bacteria; in the obtained concentrate, adding an appropriate amount of lysozyme, lywallzyme and glass beads, performing mixing, and performing lysis on cellwalls of fungal and / or bacterial cells contained in the concentrate to obtain a cell fluid; and denaturing and degrading protein components in the cell fluid to release genomic DNA to obtain a mixed solution. The method is applicable to both the bacteria and the fungi, the lywallzyme added for lysing the fungal cell walls does not affect the extraction of the bacterial genomic DNA, and the lywallzyme added for lysing the bacterial cell walls does not affect the extraction of the fungus genomic DNA; and when a laboratory technician needs to extract genomic DNA from a strain in the clinical sample, the laboratory technician does not need to distinguish whether the strain is bacteria or fungi.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a genome DNA extraction method suitable for bacteria and / or fungi. Background technique [0002] Bacteria and fungi are ubiquitous in the environment. Under certain conditions (such as accidental wounds, surgery, use of ventilators, etc.), they can enter the blood circulation system, urinary system, lower respiratory tract, and central nervous system of the human body, resulting in serious infection and even death. [0003] Since the main component of the bacterial cell wall is peptidoglycan, while the main component of the fungal cell wall is chitin, the cell wall fragmentation methods of the two are different. Therefore in the prior art, the kit for extracting bacterial genomic DNA is used to extract bacterial genomic DNA, and the kit for extracting fungal genomic DNA is used to extract fungal genomic DNA. Genomic DNA extraction methods from bacteria and fungi. [0004] In addit...

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
Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10C12Q1/6806C12R1/445C12R1/36C12R1/72C12R1/645C12R1/22C12R1/01
CPCC12N15/1017C12Q1/6806C12Q2521/531C12Q2523/308C12Q2523/32
Inventor 闫宝山王玲于大为邵育晓徐悦刘瑞鑫樊高君
Owner BEIJING BEIER BIOENG
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap