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Specific primer pair for detecting expression level of human PAK2 gene and application thereof

A specific primer pair and human detection technology, applied in the field of specific primer pairs at the expression level, can solve the problems of unsatisfactory sensitivity and specificity

Inactive Publication Date: 2016-08-03
SUZHOU GENOARRAY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Clinical detection methods generally include fecal occult blood (FOBT) test, glycosyl antigen CA199, gastrointestinal CT examination, fibrogastroscopy, sigmoidoscopy, etc., and their sensitivity and specificity are not ideal. markers are crucial

Method used

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  • Specific primer pair for detecting expression level of human PAK2 gene and application thereof
  • Specific primer pair for detecting expression level of human PAK2 gene and application thereof
  • Specific primer pair for detecting expression level of human PAK2 gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0083] Embodiment 1, the preparation of the kit for detecting the relative expression level of human PAK2 gene

[0084] 1. Synthesis of primers

[0085] The primers for detecting the relative expression of human PAK2 gene are composed of primer PAK2-F, primer PAK2-R, primer GAPDH-F and primer GAPDH-R. Each primer is a single-stranded DNA molecule, and their nucleotide sequences are as follows: Sequence 1, Sequence 2, Sequence 3 and Sequence 4 in the list are shown.

[0086] Primer PAK2-F: 5'-AATGGCAGATTGGAGTT-3' (sequence 1 in the sequence listing);

[0087] Primer PAK2-R: 5'-CGAACTTACTACCACGAA-3' (sequence 2 in the sequence listing);

[0088] Primer GAPDH-F: 5'-CATGAGAAGTATGACAACAGCCT-3' (sequence 3 in the sequence listing);

[0089] Primer GAPDH-R: 5'-AGTCCTTCCACGATACCAAAGT-3' (sequence 4 in the sequence listing).

[0090] The primers PAK2-F, PAK2-R, GAPDH-F and GAPDH-R were synthesized artificially.

[0091] 2. Preparation of kits for detecting relative expression of h...

Embodiment 2

[0093] Example 2, the method for using the kit for detecting the relative expression of human PAK2 gene prepared in Example 1

[0094] The method for using the kit for detecting the relative expression of human PAK2 gene prepared in Example 1 is as follows:

[0095] 1. Extraction of total RNA from tissues

[0096] The steps for extracting total RNA from tissues are as follows:

[0097] (1) Take an appropriate amount of fresh tissue, add 1 mL TRIzol, and grind evenly; then add 0.2 mL chloroform, shake evenly, and centrifuge at 4°C, 14,000 rpm for 10 min.

[0098] (2) After completing step (1), transfer the upper liquid phase to a new centrifuge tube, add an equal volume of isopropanol, mix well up and down, let stand at room temperature for 10 minutes, and centrifuge at 4°C and 14,000 rpm for 10 minutes.

[0099](3) After completing step (2), discard the supernatant, add 1 mL of 75% ethanol aqueous solution (prepared by mixing distilled water and absolute ethanol), gently inv...

Embodiment 3

[0115] Example 3, using the kit for detecting the relative expression of human PAK2 gene prepared in Example 1 to detect clinical specimens

[0116] Colon cancer tissues and paracancerous tissues of 8 patients clinically diagnosed with colon cancer (all volunteers who gave informed consent) were taken, and tested according to the method of using the kit in Example 2. The colon cancer tissue and paracancerous tissue of each patient need to be tested in pairs.

[0117] Each sample was repeated 3 times.

[0118] The experimental results are shown in Table 1 and Figure 4 shown. The results showed that the expression level or relative expression level of human PAK2 gene in the colon cancer tissues of 8 clinical colon cancer patients was significantly higher than that in the adjacent tissues, and the coincidence rate was 100%. Therefore, the kit prepared in Example 1 can be used for the detection or auxiliary detection of clinical colon cancer samples.

[0119] Table 1. Test re...

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Abstract

The invention discloses a specific primer pair for detecting expression level of human PAK2 gene and application thereof. The specific primer pair disclosed by the invention is composed of two primers for amplifying a specific DNA segment. The specific DNA segment has target sequences of the primer pair composed of a primer PAK2-F and a primer PAK2-R in the human genome. The primer PAK2-F is a single-strand DNA molecule disclosed as Sequence 1 in the sequence table, and the primer PAK2-R is a single-strand DNA molecule disclosed as Sequence 2 in the sequence table. The experiment proves that the specific primer pair has the advantages of high specificity, high sensitivity, short time consumption and low detection cost, is simple to operate, and is beneficial to the detection and / or assisted detection of clinical colon cancer samples.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a pair of specific primers for detecting the expression level of human PAK2 gene and its application. Background technique [0002] Colon cancer is a common malignant tumor of the digestive tract that occurs in the colon. It is more likely to occur at the junction of the rectum and the sigmoid colon. The incidence of colon cancer ranks third among gastrointestinal tumors. Clinical detection methods generally include fecal occult blood (FOBT) test, glycosyl antigen CA199, gastrointestinal CT examination, fibrogastroscopy, sigmoidoscopy, etc., and their sensitivity and specificity are not ideal. Markers are crucial. Contents of the invention [0003] The technical problem to be solved by the present invention is how to identify or assist in identifying whether the patient to be tested is a colon cancer patient. [0004] In order to solve the above technical problems, the present i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6851C12Q1/6886C12Q2600/158C12Q2600/166C12Q2531/113C12Q2545/101
Inventor 张函槊李娟
Owner SUZHOU GENOARRAY CO LTD
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