Screening reagents G418
G418 Introduction G418 is an aminoglycoside antibiotic commonly used for resistance screening in molecular genetic experiments. It produces toxins in prokaryotic and eukaryotic cells, including bacteria, yeast, plants, and mammalian cells, as well as protozoa and worms, by inhibiting the genes of transposons Tn601 and Tn5. When the neo gene is integrated into eukaryotic cell DNA, it can initiate the transcription of the sequence encoded by the neo gene into mRNA, thereby achieving efficient expression of resistance products, enabling cells to acquire resistance and grow in selective culture media containing G418. The selection characteristic of G418 has been widely applied in gene transfer, gene knockout, resistance screening, and transgenic animals. The mechanism of action of aminoglycoside antibiotics is that aminoglycoside antibiotics have a strong cytotoxic effect, mainly acting on the ribosomes of cells and inhibiting protein synthesis.The process of action is： 1. Aminoglycoside molecules adsorb onto the cell surface, causing mild damage to the cell membrane; 2. A small number of aminoglycoside molecules enter cells and bind to ribosomes, affecting the ongoing process of peptide chain elongation, causing misreading of the genetic code and abnormal protein expression; 3. Abnormal proteins bind to the cell membrane, leading to membrane leakage; 4. A large number of aminoglycoside molecules enter cells, forming more abnormal proteins, exacerbating cell membrane leakage and drug intake; 5. When the number of aminoglycoside molecules entering the bacterial cell significantly exceeds the number of ribosomes, the initial stage of protein synthesis in the ribosome is completely stopped, and this action is irreversible, leading to rapid bacterial death. As a result of the action of aminoglycoside antibiotics, important substances such as potassium ions, adenine, and nucleotides are leaked out of cells, leading to respiratory inhibition and inhibition of DNA and RNA synthesis. G418 quality indicators G418: White powder, melting point 138-144 ℃, soluble in water and methanol. CAS No.：108321-42-2 Molecular formula: C20H 40N4 O10 · 2H2 SO4 Molecular weight: 692.71
 Rotation ratio:+104 ° -+121 ° Moisture content: ≤ 10% Absorption value: ≤ 0.015 (280nm, 1mg/ml), ≤ 0.1 (570nm, 100mg/ml) Potency:>700 ug/mg Storage and transportation Transport at room temperature, store dry powder at room temperature, dissolve in water and methanol, and store the solution at -20 ℃. G418 usage method Due to the varying sensitivity of eukaryotic cell lines to G418, the amount required to kill non stably transfected cells during stable transfection of each new cell line or strain needs to be optimized through experiments. The most suitable amount is obtained by establishing a cell death curve. Dilute the cells to 1000cell/ml, add 100ul of G418 to each well in a 24 well plate with culture medium, and dilute the G418 concentration in each well to 12 levels including 0100, 0200, 300400, 500600, 700700800, 900, 1000, 1100ug/ml. Incubate for 10-14 days, using the lowest cell death concentration as the benchmark, usually around 400-800. When screening, use one level higher than this concentration, and maintain at half of the screening concentration. G418 Select concentration range
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