Establishment and synthesis method of dual-targeting DNA (deoxyribonucleic acid) nano drug-loaded complex
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A nano-drug loading and complex technology, applied in the field of biomedicine, to achieve strong resistance to nuclease degradation and important application prospects
Inactive Publication Date: 2021-11-30
QUANZHOU NORMAL UNIV
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Embodiment 1
[0050] Example 1 Materials and Methods
[0051] The cell line used in the present invention is a human malignant melanoma cell A375, purchased in the China Typical Casting Savings Center. The present invention constructs a single-stranded DNA of DNA tetrahedron, and is purchased from Shanghai's Shanghai Synthesis Department, specific sequences and modification methods and modified sites.
[0052] Table 1 DNA single-strand sequence and modification method
[0053]
[0054] The nucleic acid adapter according to the present invention has been purchased from Shanghai's Shanghai Synthesis Department, specific sequences and modified methods and modified sites below.
[0055] Table 2 Nucleic acid adapter sequence and modification method
[0056]
[0057]
[0058] 1.1 Preparation of the invention relates to a solution
[0059] 1) 10% complete cell culture medium: Take DMEM high sugar medium 180ml, FBS special fetal bovine serum 20ml, stored to high pressure sterilization and dried 2...
Embodiment 2
[0093] Example 2 Experimental Results
[0094] Construction of the first part of DNA nano-mixed complex
[0095] 1. Construction of single-targeted and double targeting DNA nano-carbal complex
[0096] 1.1DNA tetrahedron construction
[0097] In the present invention, the use of the DNA tetrahedron structure of the reported DNA is used, although the present invention adds a sticky end of the viscous end, but the design of the viscous end does not affect the DNA tetrahedral space structure. The built-in "DNA Tsui-Aptamer-AS1411" spatial structure is special, and the commonly used Marker marks are inaccurate. We increase the DNA single chain by PCR, and the increase in nucleic acid chain molecular weight during the mutual verification process, and the molecular weight change in Page glue precision is verified. As can be seen from the results, by increasing the constructed DNA single-strand, PAGE glue exhibits a strip, strips, and molecular weight exhibits a clear difference. The DNA...
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Abstract
The invention discloses an establishment and synthesis method of a dual-targeting DNA (deoxyribonucleic acid) nano drug-loaded complex. The drug-loaded complex is prepared by the following steps: simultaneously connecting an Aptamer 1 and a natural AS1411 or a chemically modified AS1411 to a DNA tetrahedron in a manner of complementary base pairing, so as to respectively obtain a DNA tetrahedral Aptamer 1-AS1411, a DNA tetrahedral Aptamer 1-AS1411-1, a DNA tetrahedral Aptamer 1-AS1411-2 and a DNA tetrahedral Aptamer 1-AS1411-3. The drug-loaded complex is applied to preparation of drugs for treating melanoma. According to the invention, four dual-targeting DNA nano drug-loaded systems based on CD44 and nucleolin nucleic acid aptamers are constructed, and compared with a single-targeting drug-loaded complex and a negative control group, the four dual-targeting drug-loaded complexes have stronger nuclease degradation resistance, capability of targeting melanoma A375 cells, capability of inhibiting proliferation of the A375 cells and capability of inducing the A375 cells to generate G1 phase retardation.
Description
Technical field [0001] The present invention relates to the field of biopharmaceutical, and more particularly to the establishment and synthesis of diblant DNA nano-drug complexes. Background technique [0002] Malignant Melanoma is one of the high degree of malignine in human skin tumors. In recent years, the incidence of melanoma continues to increase, and is a tendency to be young. Although the latest developments in immunotherapy and molecular targeting treatment provide some more effective treatment, these treatments still do not completely solve the problem of poisonous side effects of drugs. Therefore, developing a new target for melanoma, reducing the deposits of chemotherapeutic drugs, and is critical to improving overall efficacy of melanoma treatment. [0003] CD44 molecules are a class of transmembrane monocloproteins, which mediates a variety of biological functions of cells between cells and cells, cells and extracellular matrices. The study found that CD44 has been...
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