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Plant transgenic method based on silica nanoparticles as gene carrier

A nanoparticle, silicon dioxide technology, applied in the fields of botanical equipment and methods, angiosperms/flowering plants, introduction of foreign genetic material using vectors, etc., can solve problems such as unreported, achieve low price, improve transformation efficiency, Easy to prepare effect

Inactive Publication Date: 2011-11-30
JILIN AGRICULTURAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Nanoparticles have been reported as gene carriers for plant transgenic methods (Biotechnol.J., 2008, 3, 1078-1082; Biotechnology Bulletin, 2009, 6: 75-80), but nanoparticles mediate exogenous Stable expression of the target gene transferred into plants has not been reported so far

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Biological Modification of Silica Nanoparticles of Different Sizes

[0026] (1) Accurately weigh 5 mg of 3nm, 50nm, 100nm, 500nm, and 1000nm silica nanoparticles into a centrifuge tube, add 5mL of sodium phosphate buffer solution with pH=7.2, and place in a 300W, 40KHz ultrasonic instrument for ultrasonic dispersion 30-60min, control the temperature below 37°C during the ultrasonic treatment. After sonication, the solution was uniform and milky.

[0027] (2) Take the 3nm, 50nm, 100nm, 500nm, 1000nm 1mg / mL silicon dioxide nanoparticle solution dispersed by ultrasound, respectively, and the PLL of 1mg / mL according to the mass ratio of 20:1, 10:1, 5:1 Mix according to the ratio, mix well by inverting up and down, and let stand at room temperature for 1 hour. The surface charge of the PLL-modified silica nanoparticles was analyzed by a zeta potential instrument. The results show that the zeta potential value is the highest when the mass of the above various silica nanopa...

Embodiment 2

[0029] Preparation of complexes of silica nanoparticles with genes in different sizes

[0030]Take respectively the 3nm, 50nm, 100nm, 500nm, 1000nm silicon dioxide nanoparticles modified by PLL with the mass ratio of silicon dioxide nanoparticles and PLL prepared by the method in Example 1 being 10:1, and the target gene at 30:1, The mass ratio of 20:1, 10:1, and 5:1 was mixed, and left at room temperature for 0.5-1 hour to prepare the silica nanoparticle-gene complex. 1% agarose gel was used to detect the combination of silica nanoparticles with different sizes and genes. The results show that silica nanoparticles can effectively bind to DNA molecules.

Embodiment 3

[0032] Analysis of Gene Protection by Silica Nanoparticles

[0033] Take the 3nm, 50nm, 100nm silicon dioxide nanoparticles modified by PLL prepared in Example 2 and the silicon dioxide nanoparticles prepared by the mass ratio of 30:1, 20:1, 10:1, 5:1- The gene complex was subjected to ultrasonic treatment, and the target gene not combined with silica nanoparticles was used as a blank control, and was subjected to ultrasonic treatment for 20 min, 30 min, and 40 min in an ultrasonic instrument with parameters of 60W and 40KHz, respectively. 1% agarose gel detection. The results showed that silica nanoparticles of various sizes could effectively protect genes from ultrasonic damage, while genes not combined with silica nanoparticles were destroyed under the above three different conditions.

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Abstract

A plant transgenic method based on silica nanoparticles as a gene carrier includes gene gun and ultrasonic transformation methods. Silicon nanoparticles of different sizes are surface-modified with polylysine (PLL) to construct nano-gene carriers. The modified nanoparticles can effectively combine with genes through electrostatic interaction to form silica nanoparticles-gene complexes , respectively transformed plant receptor materials by gene gun method and ultrasonic method, and successfully realized the efficient and stable expression of exogenous genes in plant cells. The method of the invention has the advantages of simple operation, low price, high conversion efficiency, and the conversion material is not limited by plant species type and tissue organ type. The method of the invention lays a good foundation for the wide application of the silicon dioxide nanometer gene carrier in plant transgenesis.

Description

technical field [0001] The invention relates to a method for using silicon dioxide nanoparticles as a gene carrier to mediate exogenous gene transformation into plant materials and obtain stable expression, and belongs to the field of nanobiotechnology. Background technique [0002] The emergence and development of nanotechnology has injected new vitality into the development of life sciences. With the application of nanobiotechnology in the field of animal and human medicine, nanoparticles as a new type of gene carrier have also received more and more attention. However, There are few reports on the application of nanoparticles as gene carriers in plant transgenics. At present, nano-gene carriers are mainly divided into three categories: inorganic nanoparticles, high molecular polymers and chitosan. Silica nanoparticles are an important inorganic nanoparticle, which has been applied in the research of biomedical fields such as drug sustained release, gene carrier and biose...

Claims

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Application Information

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IPC IPC(8): C12N15/82A01H4/00A01H5/00
Inventor 付玉芹王丕武李鲁华吕长利
Owner JILIN AGRICULTURAL UNIV
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