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Rapid screening of tissue sample of rectal cancer aptamer and application of rectal cancer aptamer to preparation of detection preparation

A nucleic acid aptamer and rectal cancer technology, applied in DNA preparation, measurement devices, recombinant DNA technology, etc., to achieve low immunogenicity, good stability, and strong tissue permeability

Inactive Publication Date: 2021-05-07
HUNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods still have certain deficiencies in the detection of early rectal cancer without obvious symptoms.

Method used

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  • Rapid screening of tissue sample of rectal cancer aptamer and application of rectal cancer aptamer to preparation of detection preparation
  • Rapid screening of tissue sample of rectal cancer aptamer and application of rectal cancer aptamer to preparation of detection preparation
  • Rapid screening of tissue sample of rectal cancer aptamer and application of rectal cancer aptamer to preparation of detection preparation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023]Example 1: Clinical rectal cancer tissue nucleic acid spiral screening

[0024]Based on Tissue-SELEX technology, the clinical rectal cancer tissue is a sieve sample, and the rectal cancer carcinoma is screened for negative sieve samples. First, the synthesis of the pro-group modification X-Aptamer library was synthesized, and then the initial library was incubated with rectal cancer carcinoma, isolated and retained and retained and retained by nucleic acid sequences bound to the cancer tissue as the first round of negative sieve libraries. At the same time, the non-combined nucleic acid sequence is incubated with clinical rectal cancer tissue, and then separated and retain the nucleic acid sequence combined with rectal cancer tissue as the first round of sieve library. Then, in the second round of screening, the positive sieve retained in the previous round is divided into three groups in sequential and sterilized water (blank control group), rectal cancer (experiment group), rec...

Embodiment 2

[0028]Example 2: Determining the strongest sequence of SW1463 cell line specific binding ability

[0029]First, pretreatment of 8 candidate sequences and control library chains. The 8 nucleic acid aptamer sequences and the powder of the control library chain were centrifuged, and the sterile deionized water was diluted to a final concentration of 250 nm, degenerate in a metal bath in a 95 ° C, 10 min, and then placed Temperature temperature. SW1463 cells were simultaneously pretreated. SW1463 was washed twice with DPBS and 0.2% EDTA was added to 37 ° C. After washing with DPBS twice, cells were collected by wash buffer (WashingBuffer, 0.45% glucose, 5 mM magnesium chloride). After centrifugation, the binding buffer (BindingBuffer: DPBS, 0.45% glucose, 5 mM magnesium chloride, 100 mg / ltRNA, 1 g / lbsa) and after treatment were added and the derived nucleic acid body sequence and library. After mixing and mixing, it was placed at 4 ° C shaker for 1 hour. After the incubation, centrifug...

Embodiment 3

[0034]Example 3: Nucleic acid body OHL-1 combined with rectal cancer cells and weakness

[0035]The nucleic acid and body OHL-1 is further studied to further study the binding progents and strength of rectal cancer cells. The OHL-1 is first prepared. We set OHL-1 to different concentration gradients: 0 nm, 100 nm, 200 nm, 400 nm, 600 nm, 800 nm, 1000 nm, 1200 nm, 1400 nm, 1600 nm. High temperature denaturation at 95 ° C for 5 min, 10 min at 4 ° C for 10 min, and it was placed in normal temperature. Second, SW1463 cells were preprocessed. The original medium in the cell culture dish was discarded, and DPBS was added 2-3 times. Digestion 1min was digested with 0.2% EDTA to digest the SW1463 cells of the adhesive state from the culture dish, and then blew the cells by washing buffer (Washing Buffer, 5 mm magnesium chloride) and collected from the centrifuge tube. . Finally, oscillating mixing is placed at 4 ° C shaker incubation for 1 h. After incubation, then centrifuge with washing buff...

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Abstract

The invention discloses a nucleic acid aptamer which can be used for detecting early rectal cancer tissue and application of the nucleic acid aptamer to clinical diagnosis and treatment of rectal cancer. The sequence of the nucleic acid aptamer is selected from any one of sequences shown in SEQ ID NO.1-SEQ ID NO.8, or the nucleic acid aptamer is subjected to label modification or transformation. The nucleic acid aptamer disclosed by the invention has similar or even more superior characteristics to a protein antibody, and the characteristics are mainly reflected in that the nucleic acid aptamer has specificity, high affinity, low toxicity and low immunogenicity, can be synthesized in vitro and modified, is convenient to store and transport, and the like. When the nucleic acid aptamer disclosed by the invention is used for early rectal cancer tissue detection after being labeled by a fluorophore, the operation is simple and easy to implement, and the detection result is more visual. Meanwhile, the nucleic acid aptamer disclosed by the invention has a short-chain nucleic acid sequence, the in-vitro synthesis cost of the nucleic acid aptamer is low, the period is short, and the reproducibility is good.

Description

Technical field[0001]The present invention relates to a nucleic acid aptamer and its applications, particularly to a nucleic acid substantial body that can be used in rectal cancer cells and early clinical rectal cancer tissue samples and its preparation test reagent.Background technique[0002]The colorectal cancer has become a malignant tumor of the third and mortality of the global incidence, of which rectal cancer accounts for 30% of all diagnosed colorectal cancer, and the male incidence is slightly higher than women. Unlike the Western countries, China's rectal cancer is higher than colon cancer. In 2014, the national tumor registration area is 13.64 / 100,000, and the median of rectal cancer is in the middle of the age of 45, in the young people The incidence is a trend in an increase. Rectal cancer refers to cancer between the junction of the toothed line to the intestinal squeezidal colon, which is one of the most common malignant tumors in the digestive tract. Although recta...

Claims

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Application Information

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IPC IPC(8): C12N15/115C12N15/10G01N33/574
CPCC12N15/1048C12N15/115C12N2310/16G01N33/57446
Inventor 胡小晓谭蔚泓欧海龙刘匀毅
Owner HUNAN UNIV
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