Agarose powder is an organic substance with chemical formula c24h38o19. It is a white or yellow bead like gel particle or powder, which is a linear polymer, and the basic structure is 1,3 linked β- D-galactose and 1,4-linked 3,6-endoether-l-galactose are alternately linked.
Agar pectin is a heterogeneous mixture composed of many smaller molecules. Agarose is generally heated to above 90 ℃ to dissolve in water, and when the temperature drops to 35-40 ℃, it forms a good semi-solid gel, which is the main feature and basis of its various uses. The performance of agarose gel is usually expressed by the strength of gel. The higher the strength, the better the performance of gel.
Agarose powder is derived from the polysaccharides of red algae, and its main component is mostly lactose, of which about 70% is agarose and 30% is branched chain agarose.
Agarose has a linear structure, with repeated disaccharide units formed alternately by D-galactose and 3,6-dehydrated galactose beta-1,4 and alpha-1,3.
Extract branched agarose from β -1, with three bonds separating another chain. The substance extracted from seaweed with hot water contains about 40% agar.
Commercial agar (usually referred to as agar) is usually in the form of thin sheets or loose ropes and is commonly used as an edible chewing gum, drug packaging agent, or bacterial culture medium.
Pure agarose is often used as a semi-solid support in biochemical laboratories, for electrophoresis, chromatography, and other techniques, for the separation and analysis of biomolecules or small molecules.
Agarose Powder is a polysaccharide extracted from seaweed and is the main component of agar. Since its discovery, it has been widely used in many fields due to its unique physical and chemical properties, such as gel forming ability, thermal stability, chemical stability and biocompatibility. The following is a detailed explanation of its purpose:
Application in Biomedical Research
Agarose gel electrophoresis is one of the most commonly used separation and analysis techniques in molecular biology. As the medium of gel electrophoresis, agarose has the advantages of large pore size, wide separation range and simple operation. In the process of electrophoresis, biological macromolecules such as DNA, RNA or protein migrate through the pores of agarose gel under the action of electric field. Due to the differences in charge, size and shape of different molecules, their migration speed in gel is different, thus achieving separation.
Agarose gel electrophoresis is widely used in gene cloning, gene expression analysis and gene mutation detection. It is commonly used as a support or matrix in cell culture. Due to its excellent biocompatibility and chemical stability, agarose can provide a stable growth environment for cells. For example, in the soft agar colony formation experiment, agarose is used to prepare semi-solid culture medium to detect the colony formation ability of cells, which is of great significance for evaluating the degree of malignant transformation of cells.
In addition, agarose can also be combined with other biomaterials to prepare composite scaffold materials for use in tissue engineering and regenerative medicine. Agarose gel also plays an important role in immunodiffusion and immunoelectrophoresis. In these technologies, agarose gel, as the medium of antigen and antibody reaction, can form clear precipitation lines or rings, which can be used to detect and analyze the presence and concentration of antigen or antibody. The pore size and charge property of agarose gel can affect the diffusion rate and binding efficiency of antigen and antibody, so the experimental conditions can be optimized by adjusting the concentration and type of agarose.
Application in Molecular Biology Experiments
Agarose gel electrophoresis can not only be used to separate DNA and RNA, but also be combined with other technologies to achieve their purification. For example, in the process of DNA extraction, agarose gel electrophoresis can be used to detect the integrity and purity of DNA, and then the target DNA segment can be separated from the gel by gel cutting and recovery. This method has the advantages of easy operation and high recovery rate. In nucleic acid hybridization experiments, agarose gel can be used as a solid phase support to fix DNA or RNA probes on the gel, and then hybridize with the labeled target nucleic acid.
By detecting the presence and intensity of hybridization signals, the presence and concentration of target nucleic acids can be determined. The pore size and charge property of agarose gel can affect the binding efficiency of probe and target nucleic acid, so the hybridization conditions can be optimized by adjusting the concentration and type of agarose. Polymerase chain reaction (PCR) is one of the most commonly used techniques in molecular biology for amplifying DNA. In the analysis of PCR products, agarose gel electrophoresis is one of the most commonly used methods. By separating PCR products through electrophoresis, the size and purity of the amplified fragments can be detected, thereby determining whether the PCR reaction is successful and whether there are non-specific amplification issues.
Application in Biochemical Analysis
Although the pore size of agarose gel is relatively large, it is not suitable for separating small molecular proteins, but it can still be used for protein separation and purification in some cases. For example, in preparative gel electrophoresis, agarose gel can be used to separate and purify proteins with larger molecular weights.
In addition, agarose can also be combined with other materials to prepare composite gel to expand its separation range and improve the separation efficiency. In some enzyme activity determination experiments, agarose gel can be used as a carrier for enzyme immobilization. Immobilization of the enzyme on agarose gel can maintain the activity and stability of the enzyme and facilitate the separation and recovery of the enzyme from the reactants. This method has wide applications in the field of enzyme engineering.
Application in Microbiology
Agarose is commonly used as a coagulant in microbial culture media. Compared with agar, agarose has higher purity and lower impurity content, making it more suitable for microbial cultivation. When preparing solid culture medium, dissolve agarose in hot water, then add other nutrients and antibiotics, and cool to form solid culture medium.
This type of culture medium can provide a stable growth environment for microorganisms, facilitating their isolation, purification, and identification. In microbiological identification experiments, agarose gel can be used to prepare specific identification media. For example, when identifying intestinal bacteria, agarose culture medium containing specific sugars can be used to distinguish different types of bacteria by observing their fermentation of sugars. In addition, agarose gel can also be used to prepare biochemical reaction media for microorganisms to detect their metabolic characteristics and enzyme activities.
Agarose production process can be divided into four parts: pretreatment, extraction, DEAE cellulose treatment and dehydration.
The pretreated powdery raw materials are directly dried at 80C for 1 / 2 h. the strip and block raw materials are washed with water, cut into small pieces, and baked in.
Extraction: after stirring and extracting with acetic acid sodium acetate mixed ester solution for several hours, remove water by suction, wash, then stir and extract with sodium carbonate sodium bicarbonate buffer solution for several hours, and then pump, filter and wash with water.
The agar gel extracted by the buffer solution is dissolved into 4% solution by heating with water, and then added with DEAE cellulose for stirring for 3-4 h under heat preservation, and then centrifuged. The filtrate is collected and filtered to obtain a white solution in colloidal state.
Dehydration: after heating the white filtrate, add 95% ethanol rapidly while stirring until there is no obvious precipitation, cool it to room temperature, pour out the upper layer solution, wash and dry the precipitate to obtain the product, with a yield of 71%.
Reagent recovery DEAE cellulose recovery: after centrifugation, the precipitate is boiled with water, filtered on 100 mesh silk cloth, repeated several times until the filtrate is clear, then washed with 0.5mol/1 HCI, 0.5mol/1 NaOH and water, and dried. The recovery rate is 95% The 80 C fraction was collected by direct distillation of ethanolic aqueous solution recovered from Yichun aqueous solution Then add solid sodium chloride to saturation and continue to distill. 70% can be charged. After the sodium chloride in the bottle is evaporated, it can also be charged by valve.
Agarose powder is usually heated to above 90 ° C to dissolve in water. When the temperature drops to 35-40 ° C, it will form a good semi-solid gel, which is also its main feature and the basis for various uses. The performance of agarose gel is usually expressed by the strength of gel. The higher the concentration, the better the effect of gel. The gel of agarose is caused by the existence of hydrogen bond. Any factor that may destroy the hydrogen bond may lead to the destruction of gel. Agarose is hydrophilic and almost completely free of charged groups. It rarely causes denaturation and adsorption to sensitive biomolecules. It is an ideal inert carrier.
During the preparation of agarose, it is necessary to remove agarose pectin as much as possible, otherwise there may be a very small amount of sulfate and pyruvate replacing the ionized groups in agarose, which can lead to electroosmosis (EEO) and affect particle movement. The sulfate content of high-quality agarose is relatively low, usually below 0.2%, and its electroosmotic effect is relatively small, usually below 0.13. That's why agar is much more expensive than agar.
In 1937, Araki first isolated agar from agar.
It was not until 1961 that hjertin first discovered the excellent properties of agarose, which attracted increasing attention and began industrial production. Agarose is widely used in clinical testing, biochemical analysis, and separation of biomolecules.
Agarose powder is a basic tool in molecular biology and biotechnology. It is widely used in gel electrophoresis, affinity chromatography, immunodiffusion, immunoelectrophoresis and other fields.
Its unique characteristics, such as ease of use, multifunctionality, low toxicity, and cost-effectiveness, make it an indispensable component in laboratory research.
Despite the limitations of limited resolution and brittleness, the latest advances in agar gel technology, including high-resolution gel, prefabricated gel and agar based microfluidic devices, have expanded their practicality and improved their performance.
With the continuous progress of scientific research, agarose powder will undoubtedly continue to be a valuable and essential reagent in the toolbox of molecular biology and biotechnology.
FAQ
What is agarose and what is its purpose?
Agarose gel electrophoresis has proven to be an efficient and effective way of separating nucleic acids. Agarose's high gel strength allows for the handling of low percentage gels for the separation of large DNA fragments.
What is agarose and agar?
Agarose is a polysaccharide extracted from seaweeds (e.g. red algae) and its main component is polygalactose. Agar consists of agarose and agar pectin together and is purified to obtain agarose. Agarose is a linear polymorph, while agar pectin is a heterogeneous mixture of many smaller molecules.
What is the meaning of agarose?
Agarose is defined as a high-molecular-weight polysaccharide extracted from marine red algae, specifically agarocytes of Rhodophyceae, that forms gels ideal for cellular studies and gel electrophoresis due to its biocompatibility and thermo-reversible gelation properties.
What is the main purpose of agarose powder?
Used mainly for gel electrophoresis, agarose has other applications such as gel filtration chromatography and affinity purification. Its most common use is in the preparation of agarose gels, where pore size depends on agarose concentration.
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