Composition and method for detecting group A streptococcus,

A composition, streptococcal technology, applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., can solve the problem of unsatisfactory sensitivity

Active Publication Date: 2020-04-17
SANSURE BIOTECH INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Chinese patent publication CN107058622A discloses a kit for the joint detection of respiratory pathogens by multiplex fluorescent PCR method, which includes GAS-specific primers and probes, but the sensitivity in detecting GAS is unsatisfactory, and group B streptococcus is not used for detection of respiratory pathogens. Specificity is verified

Method used

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  • Composition and method for detecting group A streptococcus,
  • Composition and method for detecting group A streptococcus,
  • Composition and method for detecting group A streptococcus,

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0075] Example 1 is designed to detect primers and probes for GAS

[0076] According to the spe B gene sequence of GAS, a total of 10 upstream primers, downstream primers and probe combinations were designed. Table 1 below takes three groups as examples to show the specific sequences and detection results of each primer and probe.

[0077] Table 1 Example of primer probe sequence based on GAS-based spe B gene sequence design

[0078]

Embodiment 2

[0079] Embodiment 2 detects GAS in the sample

[0080] 2.1 Extraction of DNA from GAS

[0081] Use the magnetic bead method nucleic acid extraction or purification reagent produced by Shengxiang Biotechnology Co., Ltd. (Xiangchang Machinery Equipment 20150021), and follow the manufacturer's instructions to extract the nucleic acid of GAS in the sample.

[0082] 2.2 Prepare PCR reaction reagents

[0083] 2.2.1 Experimental materials

[0084] Internal standard template: a recombinant of a 132 base pair artificially synthesized DNA sequence inserted into the pMD18T vector, wherein the sequence of 132 base pairs is shown in SEQ ID NO:4:

[0085] Internal standard upstream primer: as shown in SEQ ID NO:5;

[0086] Internal standard downstream primer: as shown in SEQ ID NO:6;

[0087] Internal standard probe: 5'HEX-CGCGCTTCTGGAAAGGCTTCTCAGT-BHQ1-3';

[0088] Positive control: calibrate the plasmid with known concentration, the concentration is 5.00E+05copies / ml;

[0089] Negat...

Embodiment 3

[0101] Embodiment 3 compares the detection effect of different combinations

[0102] Using combinations 1 to 3 in Example 1, 5 cases of GAS clinically positive samples were detected according to the method in Example 2, wherein the fluorescent quantitative PCR instrument was Hongshi SLAN-96, and the results were respectively: Figure 1 ~ Figure 3 The mean Ct values ​​are summarized in Table 3, as indicated.

[0103] Table 3 Summary of test results of combinations 1 to 3

[0104]

[0105] Such as Figure 1~3 As shown in Table 3, the Ct value of combination 1 is higher, indicating that its amplification efficiency and sensitivity are higher.

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Abstract

The invention relates to a composition for detecting group A streptococcus. The composition comprises a first primer with a sequence of 5'-TTAGCATTAGGTGGATTTGTTCTT-3', a second primer with a sequenceof 5'-GCTATCTTTTGCTTCTTTTTCGTTA-3'and a probe with a sequence of 5'-TAACCCAGTATTTGCCGATCAAAAC TTTGC-3'. Therefore, the group A streptococcus can be accurately detected. In addition, the invention alsoprovides a method for detecting the group A streptococcus and application thereof.

Description

technical field [0001] The invention relates to the field of in vitro diagnosis, in particular to a composition for detecting group A streptococcus. Background technique [0002] Group A streptococcus (Group A Streptococcus, GAS), which usually exhibits β-hemolysis when cultured on a blood plate, is therefore also called group A β-haemolytic streptococcus (group A β-haemolytic streptococcus), is a human Common pathogenic bacteria. The carrier rate of GAS in healthy people is 5%-15% or lower. GAS infection may be an influencing factor of acute angina or skin infection, and may also be one of the causes of various purulent and non-purulent complications. According to statistics, there are 616 million cases of acute angina and 111 million cases of skin infection caused by GAS every year in the world. There are about 18.1 million cases of GAS-related serious diseases, and the incidence rate is increasing at an annual rate of about 1.78 million cases, and at least 517,000 peop...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/689C12Q1/6851C12Q1/14C12N15/11C12R1/46
CPCC12Q1/689C12Q1/6851C12Q2600/166C12Q2531/113C12Q2563/107C12Q2545/101C12Q2545/113Y02A50/30
Inventor 易晓明黄河宴丝妮王铁军戴立忠邓中平刘佳
Owner SANSURE BIOTECH INC
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