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Kit for detecting staphylococcus epidermidis and application of kit

A technology for Staphylococcus epidermidis and detection kits, which can be used in microorganism-based methods, microbial determination/inspection, biochemical equipment and methods, etc., and can solve problems such as low accuracy, low detection rate, and poor repeatability.

Active Publication Date: 2015-04-22
BIOSINO BIO TECH & SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The traditional PCR detection method of Staphylococcus epidermidis has the disadvantages of long time-consuming, low detection rate, low accuracy and poor repeatability

Method used

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  • Kit for detecting staphylococcus epidermidis and application of kit
  • Kit for detecting staphylococcus epidermidis and application of kit
  • Kit for detecting staphylococcus epidermidis and application of kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] The design of embodiment 1 primer

[0023] The nucleotide sequence of detection primer provided by the invention:

[0024] Forward primer F: 5'-GTTGCGGAATCATGGTACTGT-3';

[0025] Reverse primer R: 5'-AGACAGTTACTCTCACAGGCA-3'.

Embodiment 2

[0026] Embodiment 2 Establishment of PCR detection method

[0027] 1. Extract template DNA

[0028] Boiling method to extract template DNA: take 1mL concentration of 10 6 Place the Staphylococcus epidermidis bacterial liquid of McFarland units in a 1.5mL sterile centrifuge tube; centrifuge at 10000rpm at room temperature for 3min, discard the supernatant, and add 80μL of cell lysate (1% Triton, 10mM Tris-HCL, 1mM pH 8.0 EDTA), boiled for 6 minutes, then centrifuged at 10,000 rpm for 3 minutes at room temperature, and the supernatant was taken and placed at 4°C for later use or stored at -20°C.

[0029] 2. PCR reaction system optimization test (25 μL)

[0030] System one:

[0031]

[0032] System two:

[0033]

[0034] System three:

[0035]

[0036] Depend on figure 1 It can be seen that in the reaction system 1, when 1.5 μL of the upstream and downstream primers are added respectively (the final concentration is 0.6 μM), false positives appear in the E. coli te...

Embodiment 3

[0042] Embodiment 3 specificity test

[0043] The template DNA is selected from: sterile water, human genomic DNA, Enterococcus faecium (E.faecium), Enterococcus faecalis (E.faecalis), Staphylococcus aureus (S.aureus), Staphylococcus epidermidis (S.epidemidis), Escherichia coli (E. coligeneric), Acinetobacter baumannii (A. baumannii), Enterobacter cloacae (E. coli), Klebsiella pneumoniae (K. pneumoniae) and Pseudomonas aeruginosa (P. aeruginosa).

[0044] Adopt the method of embodiment 2 to test, the result is as follows image 3 . Depend on image 3 It can be seen that only Staphylococcus epidermidis has a single band at 190bp, and no amplified band appears in other strains. It shows that the primers designed in the present invention have good specificity and can be used to detect Staphylococcus epidermidis.

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Abstract

The invention provides a kit for detecting staphylococcus epidermidis and an application of the kit, and belongs to the technical field of gene detection. According to the kit provided by the invention, PCR amplification is carried out on a to-be-detected sample by virtue of primers in SEQ ID NO.1-2; the amplification products are subjected to agarose gel electrophoresis detection; if a single strip exists at the position of 190bp, the result is positive; and or else, the result is negative. The primers disclosed by the invention are good in specificity; the detection method is fast, simple, high in accuracy, and good in sensitivity; and a novel method is provided for detection of the staphylococcus epidermidis.

Description

technical field [0001] The invention relates to the technical field of gene detection, in particular to a kit for detecting Staphylococcus epidermidis and its application. Background technique [0002] Staphylococcus epidermidis is a common bacterium that grows on the epidermis of organisms. It is parasitic on the skin, vagina and other parts of the human body, and belongs to the normal flora type. Staphylococci are Gram-positive bacteria belonging to the family Bacteriaceae and the genus Staphylococcus. After senescence, death, or phagocytosis by white blood cells, or certain drug-resistant strains can be stained Gram-negative. Staphylococcus epidermidis is widely distributed in nature, most of which are non-pathogenic bacteria, and only a few can cause suppurative infections in humans and animals. Staphylococcus is the most common pyogenic coccus, which is an important source of cross-infection in hospitals. Staphylococcus epidermidis mainly causes carbuncle, furuncle,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/14C12N15/11C12R1/45
CPCC12Q1/689
Inventor 何犇易翔及思莹黄茜杨君
Owner BIOSINO BIO TECH & SCI