Constructing method of central system infection pathogen detection library based on high-throughput sequencing, detection method and kit

Abstract

The invention discloses a constructing method of a central system infection pathogen detection library based on high-throughput sequencing, a detection method and a kit. The library construction method comprises the following steps: extracting deoxyribonucleic acid (DNA) from a sample, and fragmenting the DNA to obtain a segmented DNA sequence; adding the segmented DNA sequence into a tail end repairing system to carry out a tail end repairing reaction, directly adding a linker sequence, a connecting enzyme, adenosine triphosphate (ATP) and polyethylene glycol into the reaction system after the tail end repairing reaction is completed, taking the ATP as an auxiliary enzyme under the action of the connecting enzyme to catalyze the linker sequence to be connected to two ends of the product of the tail end repairing reaction, so that a linker connecting product is obtained; and purifying the linker connecting product, and adding a nucleic acid single chain which is complementary to two ends of the linker sequence and used as a primer for polymerase chain reaction (PCR) amplification. The method can be used to realize rapid and broad-spectrum central system infection pathogen microorganism detection, too much pre-known clinical information of detection samples is not needed, detection can be carried out quickly and efficiently, the detection result is free of deviation, and a problem that the library construction failure rate of low-concentration cerebrospinal fluid samples is high can be solved.

Description

technical field [0001] The invention relates to the technical field of microbial detection, in particular to a high-throughput sequencing-based method for constructing a central system infection pathogen detection library, a detection method and a kit. Background technique [0002] Acute encephalitis and meningitis and other central system infection syndromes include a series of acute inflammatory diseases caused by various pathogens and characterized by brain parenchyma and / or meningeal infection. It is a disease that seriously threatens human health. According to the World Health Organization, the global incidence of viral encephalitis is about 3.5 / 100,000-7.4 / 100,000. About 125,000 infants and young children die of bacterial meningitis every year in the world, and the disability rate of bacterial meningitis is as high as 25%-50%. Early and effective diagnosis of the causative agent of encephalitic meningitis syndrome can effectively avoid poor prognosis and even death du...

AI Technical Summary

Problems solved by technology

[0005] However, microbial analysis based on high-throughput sequencing currently has no solution for clinical testing needs. The methods used in scientific research are costly, time-consuming, and require high sample input volume, which cannot meet clinical needs.

Taking cerebrospinal fluid samples as starting materials as an example, the nucleic acid concentration of cerebrospinal fluid samples varies greatly, resulting in a high failure rate of sequencing library construction

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