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Nucleic acid reagent, kit, system and method for detecting invasive fungi

An invasive fungus and kit technology, applied in the biological field, can solve the problems of complex and cumbersome test procedures, increase the complexity of operations, etc., and achieve the effect of overcoming time-consuming and saving time.

Inactive Publication Date: 2019-12-10
北京卓诚惠生生物科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

PCR detection requires complex sample processing, cumbersome test procedures and complex result judgments
In addition, the detection targets that RT-PCR can cover are limited, and multi-system settings are required to achieve multi-target detection, which increases the complexity of the operation

Method used

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  • Nucleic acid reagent, kit, system and method for detecting invasive fungi
  • Nucleic acid reagent, kit, system and method for detecting invasive fungi
  • Nucleic acid reagent, kit, system and method for detecting invasive fungi

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] Embodiment 1 detection method and detection result judgment

[0074] 1. Primer and probe synthesis

[0075] Sequence synthesis was performed according to the primer sequences shown in Table 1 and the probe sequences shown in Table 2. In the probe, FAM is 6-carboxyfluorescein, CY5 is 5H-indocyanine, ROX is 6-carboxy-X-rhodamine, VIC is a dye purchased from ABI, and BHQ1-3 is a quenching group.

[0076] Table 1

[0077]

[0078] Table 2

[0079] detection target probe code probe sequence SEQ ID NO CA CA-P FAM-acgtggtggacgttaccgcc-BHQ1 17 CT CT-P VIC-tgtggtggccactagcaaaat-BHQ1 18 CG CG-P CY5-tctgtctrcccagcacgc-BHQ3 19 CP CP-P ROX-caaacccgagggtttgagggagaaa-BHQ2 20 Asp Asp-P FAM-tcctcgagcgtatggggct-BHQ1 21 Muc Muc-P FAM-caacttgcgctcattggta-BHQ1 22 Cn Cn-P VIC-ctcgggttttaytacctgttggactt-BHQ1 23 Pnj Pnj-P CY5-cagacttcttgcgataagg-BHQ3 24 C-au C-au-P ROX-tgtcgttattgttacta...

Embodiment 2

[0099] Embodiment 2 minimum detection limit verification

[0100] Test samples for evaluation: Candida albicans, Candida tropicalis, Candida glabrata, Candida parapsilosis, Mucormyces, Aspergillus, Cryptococcus neoformans, Pneumocystis jirovecii in blood, sputum or alveolar lavage fluid and Candida auris, the respective genomes were extracted by pyrolysis method, and the quantitative concentration was 5×10 4 copies / mL, and then serially diluted to obtain a concentration of 5×10 3 copies / mL, 5×10 2 Copies / mL, 5×10copies / mL evaluation template.

[0101] According to the detection method of Example 1, the evaluation templates of each concentration were detected respectively, and the detection was repeated 20 times for each concentration gradient, and the average value was taken as the final detection result, as shown in Table 4.

[0102] Table 4

[0103]

[0104]

[0105] It can be seen from Table 4 that the minimum detection limit of the genome of the detection target ...

Embodiment 3

[0106] Example 3 specificity verification

[0107]Select human coronavirus (purchased from Wuhan Institute of Virology, Chinese Academy of Sciences, number 2233), cytomegalovirus (purchased from Wuhan Institute of Virology, Chinese Academy of Sciences, number 303), enterovirus (purchased from Wuhan Institute of Virology, Chinese Academy of Sciences, number 977) ), measles virus (purchased from Wuhan Institute of Virology, Chinese Academy of Sciences, number 1513), human metapneumovirus (purchased from Wuhan Institute of Virology, Chinese Academy of Sciences, number 1768), mumps virus (purchased from Wuhan Institute of Virology, Chinese Academy of Sciences, No. 3145), Rhinovirus (purchased from Wuhan Institute of Virology, Chinese Academy of Sciences, No. 2343), Bacillus pertussis (purchased from Wuhan Institute of Virology, Chinese Academy of Sciences, No. 23524), Chlamydia pneumoniae (purchased from Wuhan Institute of Virology, Chinese Academy of Sciences, No. 14127), Mycopla...

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PUM

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Abstract

The invention relates to a nucleic acid reagent, kit, system and method for detecting invasive fungi. The nucleic acid reagent comprises a primer shown in SEQ ID NO.1-14 and a probe shown in SEQ ID NO.17-26, and the primer and the probe are stored independently with each other correspondingly or mixed with each other at will. Through the primer and the probe, the nucleic acid reagent, the kit, thesystem and the method of at least 9 invasive fungi of candida albicans, candida tropicalis, candida glabrata, candida parapsilosis, mucor, aspergillus, cryptococcus neoformans, pneumocystis jeroveciand candida auris, fast, comprehensive, sensitive, specific and automatic detection result determination can be achieved, and the sensitivity, specificity, simplicity and convenience of simultaneous detection of the detection target genome are significantly improved.

Description

technical field [0001] The present disclosure relates to the field of biotechnology, in particular, to a nucleic acid reagent, kit and method for detecting invasive fungi. Background technique [0002] Invasive fungal infection (IFI) refers to the pathological changes and pathophysiological processes in which fungi invade human tissues and blood, grow and reproduce in them, leading to tissue damage, organ dysfunction and inflammatory response. In the past 30 years, with the development and utilization of solid organ and hematopoietic stem cell transplantation technology and catheter intervention and indwelling technology, as well as the extensive application of immunosuppressants, radiotherapy and chemotherapy, the morbidity and mortality of invasive fungal infection have significantly increased globally. raised. Epidemiological studies have shown that the rate of invasive fungal infection in hospitals is 7.5%, the rate of invasive fungal infection in cancer patients is 8%,...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12Q1/686C12N15/11C12Q1/04C12R1/725C12R1/74C12R1/72C12R1/785C12R1/66C12R1/645
CPCC12Q1/686C12Q1/6895C12Q2600/16C12Q2537/143C12Q2563/107
Inventor 林笑冬王晓艳王雷吴瑞倩张志强
Owner 北京卓诚惠生生物科技股份有限公司
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