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Probes for TORCH detection, gene chip, kit and detection method

A detection kit and gene chip technology, applied in the field of molecular biology, can solve problems such as weak signal, and achieve the effects of simplifying test steps, widening the market potential for clinical application, and reducing investment costs and application costs.

Pending Publication Date: 2018-07-17
重庆高圣生物医药有限责任公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Its signal is weak and must be excited to emit light, requiring expensive laser scanning equipment

Method used

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  • Probes for TORCH detection, gene chip, kit and detection method
  • Probes for TORCH detection, gene chip, kit and detection method
  • Probes for TORCH detection, gene chip, kit and detection method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] [Example 1] Design of target fragments, primers and probes

[0029] Search through the NCBI database including Toxoplasma gondii TOX, rubella virus RV, cytomegalovirus CMV, herpes simplex virus type I and herpes simplex virus type II gene sequences, and select conserved sequences as targets (Gene Bank IDs are AF179871, M15240, KX544834, MF156584 and KY363356), according to the design principles of primers and probes, the amplification primers and probes of each pathogen gene were designed. The end was labeled with biotin, the specific sequence is as follows:

[0030] Toxoplasma gondii TOX

[0031] Target sequence amplification primer pair, the sequences are respectively:

[0032] TOX-F: SEQ ID NO.11

[0033] TOX-R: SEQ ID NO. 12;

[0034] rubella virus RV

[0035] Target sequence amplification primer pair, the sequences are respectively:

[0036] RV-F: SEQ ID NO.13

[0037] RV-R: SEQ ID NO. 14;

[0038] CMV

[0039] Target sequence amplification primer pair, th...

Embodiment 2

[0050] [Example 2] Preparation of chip

[0051] Amino-modified probes and control probes were arranged according to Table 1, and the probes were respectively spotted on the NC. Two parallel spotting points were set for each group, and the probes were immobilized by ultraviolet crosslinking for 20 minutes.

[0052] Control ProbeT probe sequence SEQ ID NO.26.

[0053] Table 1 TORCH detection chip matrix

[0054]

Embodiment 3

[0055] [Example 3] Detection method

[0056] 1. Genome extraction: Use the DNA / RNA co-extraction kit from Tiangen Biochemical Technology (Beijing) Co., Ltd., Cat. No. DP422.

[0057] 2. PCR amplification: see Tables 2 and 3 for system components.

[0058] The composition of TOX, CMV, HSV I and HSV II multiplex PCR amplification system is shown in Table 2 below:

[0059] Table 2 Composition of TOX, CMV, HSV I and HSV II multiplex PCR amplification system

[0060]

[0061] The PCR reaction program was: 95°C for 5min; 32 cycles of 95°C for 10s, 58°C for 30s, 72°C for 30s; 72°C for 10min.

[0062] Rubella virus RV is a retrovirus, and the system composition is shown in Table 3 below:

[0063] Table 3 RV amplification system

[0064]

[0065] The PCR reaction program was: 10-30 min at 45°C; 5 min at 95°C; 32 cycles of 10 s at 95°C, 30 s at 58°C, 30 s at 72°C; 10 min at 72°C.

[0066] The one-step RT-PCR amplification kit (AMV) of Sangon Bioengineering (Shanghai) Co., Ltd...

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Abstract

The invention discloses a set of probes for TORCH detection. The probes comprise the following components: a Toxoplasma Gondii TOX probe 1 and 2, wherein the sequences respectively are SEQ ID No.1-2;a rubella virus RV probe 1 and 2, wherein the sequences respectively are SEQ ID No.3-4; cytomegalovirus TOX probe 1 and 2, wherein the sequences respectively are SEQ ID No.5-6; herpes simplex virus Itype HSV I probe 1 and 2, wherein the sequences respectively are SEQ ID No.7-8; herpes simplex virus II type HSV II probe 1 and 2, wherein the sequences respectively are SEQ ID No.9-10. The inventionalso discloses a gene chip, a kit and a detection method thereof containing the probes. Optical amplification of hybridization signals with visible light is carried out, signal resolution is high, andthe signals can be directly interpreted. The probes have the advantages of simple operation, low cost, and easy reading of results; clinic on-site rapid detection demands of hospital are satisfied, and the results are accurate and reliable.

Description

technical field [0001] The invention belongs to the field of molecular biology, and relates to a probe, gene chip, kit and detection method for TORCH detection. Background technique [0002] TORCH is the acronym for the causative agent of a group of diseases of perinatal infection. Andres Nahmias et al. used TORCH for the first time to represent four congenital infectious diseases affecting fetuses and newborns, namely Toxoplasma (TOX), rubella virus (Rubella virus, RV), cytomegalovirus (Cytomegalo virus, CMV) and simple Herpes virus (Herpessiplex virus I / II, HSV I / II). Over time, this abbreviation has gradually expanded. The O stands for others, mainly including syphilis, parvovirus B19, enterovirus, hepatitis B and human immunodeficiency virus (HIV). [0003] TORCH pathogens can cause a variety of clinical symptoms, the first infection of pregnant women, these pathogens can be vertically transmitted to the baby through the placenta, resulting in habitual abortion, prema...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6893C12Q1/686C12Q1/6837C12N15/11C12R1/93
CPCC12Q1/6837C12Q1/686C12Q1/6893C12Q1/701C12Q1/705C12Q2600/16C12Q2537/143
Inventor 周勇项周
Owner 重庆高圣生物医药有限责任公司
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