Real-time fluorescence quantitative PCR (Polymerase Chain Reaction) kit for detecting candida tropicalis

A Candida, fluorescence quantitative technology, applied in microorganism-based methods, microorganisms, biochemical equipment and methods, etc., can solve the problems of time-consuming, expensive, complicated operation, etc., and achieve cost reduction, easy operation, and operation. simple effect

Inactive Publication Date: 2011-09-14
SOUTHERN MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Ordinary PCR is easy to cause laboratory pollution and is rarely used in clinical testing. Universal primer PCR has the advantage of being fast and sensitive, but the specificity of separation is poor, especially for the identification of drug-resistant strains; nucleic acid probes require a large number of probes to increase specificity. Needle, which is expensive and time-consuming, cannot be used for routine laboratory examination; PCR-SSCP, PCR-RFLP, gene chip technology, etc. have good sensitivity and specificity, but the operation is relatively complicated and the requirements for equipment are high, so they are mostly used for scientific research , are not currently suitable for clinical testing

Method used

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  • Real-time fluorescence quantitative PCR (Polymerase Chain Reaction) kit for detecting candida tropicalis
  • Real-time fluorescence quantitative PCR (Polymerase Chain Reaction) kit for detecting candida tropicalis
  • Real-time fluorescence quantitative PCR (Polymerase Chain Reaction) kit for detecting candida tropicalis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1 Candida tropicalis detection kit and its use

[0036] 1. Prepare a kit including the following components: 1 tube of primer-probe mixture (25 μl / tube), 1 tube of PCR reaction solution (250 μl / tube), 1 tube of PCR enzyme system (75 μl / tube), positive quality control product (200μl / tube) 1 tube, negative quality control product (200μl / tube) 1 tube, pure H 2 O (2000μl / tube) 1 tube. Positive quantitative standard 4 tubes (10 7 copies / ml, 10 6 copies / ml, 10 5 copies / ml, 10 4 copies / ml).

[0037] 2. Collection, preservation and transportation of specimens

[0038] 2.1 Specimen types: blood, sputum, alveolar lavage fluid, etc.

[0039] 2.2 Specimen collection, storage and transportation: Take 200ul of blood, sputum, alveolar lavage fluid, etc., put them in sterile centrifuge tubes, label them well, put them in a curling container and take them to the laboratory or at low temperature (-20℃~-70℃ ℃) frozen. Dry ice is used for long-distance transport of specime...

Embodiment 2

[0050] Example 2 Application of Candida tropicalis detection kit to detect clinical samples

[0051] Select 3 cases identified as Candida tropicalis negative through the culture method, 3 cases were identified as Candida tropicalis positive specimens through the culture method, the nucleic acid extraction of the specimen, PCR amplification and result analysis steps were carried out with reference to Example 1, and at the same time Perform negative and positive quality control tests.

[0052] Test result: the amplification curve of the negative quality control product is not S-shaped (see figure 1 ), the amplification curve of the positive quality control product is an obvious S-shaped curve (see figure 2 ), both negative and positive quality control products meet the quality control requirements of the kit; the instrument draws a standard curve based on the positive quantitative standard (see image 3 ) has a good linear relationship, indicating that the amplification effic...

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Abstract

The invention discloses a real-time fluorescence quantitative PCR (Polymerase Chain Reaction) kit for detecting candida tropicalis. The kit comprises the following substances: a PCR solution, a primer and probe mixed solution, a PCR enzyme, pure H2O and a positive quantitative standard product. The kit is used for detecting nucleic acid amplification level of the candida tropicalis, so that state of candida tropicalis infection in a patient can be reflected, and the kit can be used for monitoring, preventing and controlling fungal infection and contribute to early diagnosis and treatment of diseases; and in the real-time fluorescence quantitative PCR technology, a primer and a probe are designed aiming at a specific sequence of the nucleic acid of the candida tropicalis, so that the kit has the advantages of high specificity, high flexibility, higher flux, easiness and convenience in operation and relatively reduced cost, and is suitable for detecting various specimens of most of patients. The amplification of a specimen to be detected by the kit is finished by a commercial real-time fluorescence quantitative PCR instrument; the kit is easy to operate and has low time consumption; and pollution is reduced to the greatest extent.

Description

technical field [0001] The invention relates to a kit for detecting Candida tropicalis (Candida tropicalis), in particular to a kit for detecting Candida tropicalis with real-time fluorescent quantitative polymerase chain reaction technology. Background technique [0002] In recent years, with the continuous expansion of immunocompromised populations (including malignant tumors, blood diseases, AIDS, SARS, diabetes, autoimmune diseases, severe burns and trauma, long-term drug addicts, etc.), broad-spectrum antibiotics, corticosteroids and immune Inhibitors are widely used clinically; new technologies such as organ transplantation, catheter intubation, and interventional therapy are widely used, which has sharply increased the incidence of opportunistic fungal infections and has become one of the main causes of nosocomial infection deaths. [0003] According to the data from the US National Nosocomial Infection Surveillance Center (NNIS), the deep fungal infection rate of hos...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12R1/74
Inventor 王前龙燕郑磊
Owner SOUTHERN MEDICAL UNIVERSITY
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