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PCR-specific primers, detection kits and detection methods for genes related to benign and malignant thyroid nodules

A technology for thyroid nodules and detection kits, applied in the field of molecular biology, can solve the problems of special customization of chips, poor repeatability, and complicated operations, so as to reduce the probability of being classified as malignant, reduce the operation of thyroid nodules, and improve the effect of sensitivity

Active Publication Date: 2021-08-06
GUANGDONG GENERAL HOSPITAL +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Thyroid ultrasonography and fine-needle aspiration biopsy have poor specificity and may lead to misjudgment when distinguishing follicular thyroid neoplasia from follicular thyroid carcinoma. The accuracy rate is not high, resulting in a large number of unnecessary malignant diseases treatment case
Chip-based tumor marker detection is mainly provided by the chip provider, which has problems such as high technical cost, complicated operation, low detection sensitivity, poor repeatability, narrow analysis range, and special customization of the chip.

Method used

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  • PCR-specific primers, detection kits and detection methods for genes related to benign and malignant thyroid nodules
  • PCR-specific primers, detection kits and detection methods for genes related to benign and malignant thyroid nodules
  • PCR-specific primers, detection kits and detection methods for genes related to benign and malignant thyroid nodules

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Example 1 Multiplex PCR-specific primers, detection kit and detection method for genes related to benign and malignant thyroid nodules based on high-throughput sequencing technology

[0052] 1. Design of primers

[0053] The gene sequences related to benign and malignant thyroid nodules involved in this example were selected from the UCSC (University of California Santa Cruz, University of California, Santa Cruz) database, and hotspot mutation primers were designed based on the relevant gene sequences, and the design range included benign and malignant thyroid nodules. Mutational hotspots in associated genes.

[0054] like figure 1 As shown, in this example, a total of 27 pairs of primers were designed for hotspot mutations of genes related to thyroid nodules. Wide coverage, stable structure, and multiple detection sites.

[0055] Specifically, 27 pairs of amplification primers are the amplification primer pairs of 24 genes in Table 1 as follows:

[0056] Table 1

...

Embodiment 2

[0081] Example 2 Primer Specificity Verification

[0082] Nucleic acid was extracted from peripheral blood samples (number: 1-3), fine needle puncture samples (number: 4-6), and paraffin tissue samples (number: 7-9) using the method of step S11 in Example 1, and the concentration and purity were determined. After the determination, take qualified samples and use 10mM Tris to dilute each sample to 100ng / μL, and use 1% agarose gel electrophoresis to detect the quality of each sample (the qualification standard is the same as step S11 in Example 1), and enter the group after passing the test and perform mark. Using the method of step S12 in Example 1, the above-mentioned 9 cases of qualified samples were amplified, and the sample volume was 2 μL each. After the amplified product was purified, it was detected by 1% agarose gel electrophoresis (the qualification standard was the same as in Step S13 in Example 1. Step), 9 cases of samples were detected using specific primer amplifi...

Embodiment 3

[0084] Example 3 Primer Detection Sensitivity Verification

[0085] Using the method of step S11 in Example 1, sample nucleic acids were extracted from peripheral blood samples (number: 1), fine needle aspiration samples (number: 4), and paraffin tissue samples (number: 7) that passed the quality inspection to verify the sensitivity of primer detection. The initial concentration of each sample is 100ng / μL, and it is diluted according to the concentration gradient of 5 times, 10 times and 20 times. After dilution, the concentration of each sample is 20ng / μL, 10ng / μL and 5ng / μL respectively, and the sample name and Concentration marks. The method of step S12 in Example 1 was used to amplify the above-mentioned 9 cases of qualified diluted samples, and the sample volume was 2 μL each, and the 9 cases of samples were detected using specific primer amplification and detection methods for benign and malignant thyroid nodules related genes. The control test is the same as in Example...

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Abstract

The invention discloses a PCR-specific primer, a detection kit and a detection method for genes related to benign and malignant thyroid nodules. The sequences of the specific primers for PCR of the genes related to benign and malignant thyroid nodules of the present invention are shown in SEQ ID NO: 1 to SEQ ID NO: 54. The benign and malignant thyroid nodules related genes disclosed in the present invention are closely related to the pathological indicators of benign thyroid nodules and malignant thyroid nodules, and can be used as biomarkers to classify thyroid nodules into benign and malignant thyroid nodules. Individualized intervention treatment provides an effective basis. The PCR-specific primers, detection kit and detection method of the present invention can simultaneously detect a plurality of genes related to benign and malignant thyroid nodules in multiple samples, effectively improve detection efficiency and accuracy, reduce costs, and simplify operation steps.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a PCR-specific primer, a detection kit and a detection method for benign and malignant thyroid nodule related genes. Background technique [0002] Thyroid cancer is a common malignant tumor originating from the thyroid gland, which occupies the first place in the incidence of head and neck tumors, and its incidence has been increasing in recent years. A study in 2012 found that the incidence of thyroid cancer in women has jumped to the fifth place in women's prone to cancer. According to the data of NCI's SSER database, the incidence of thyroid cancer in 1975 was 4.04 / 100,000 population, and the incidence increased in 2007. to 11.99 / 100,000 population. [0003] At present, overtreatment of thyroid nodules is common. According to a thyroid surgery statistics in Changchun, 9216 cases of thyroid nodules treated by surgery only accounted for about 10% of malignant tumors, while ben...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6886C12Q1/6883C12Q1/6869C12N15/10C12N15/11
CPCC12N15/1013C12Q1/6869C12Q1/6883C12Q1/6886C12Q2600/156C12Q2535/122
Inventor 邝建朱瑞娟赖水青朱奇陈志江梁雅俊王龙
Owner GUANGDONG GENERAL HOSPITAL
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