Geneticin, G-418 Sulfate is a class of aminoglycoside antibiotics, whose structure is similar to neomycin kanamycin. It can interfere with the function of 80S ribosomes in eukaryotic cells, thereby blocking protein synthesis and leading to eukaryotic cell death[1,4]. Geneticin, G-418 Sulfate from 1 to 300?μg/ml is often used for clonal selection in prokaryotes and eukaryotes. Antibiotic resistance assay of clinically isolated bacterial strains found that all strains carrying 3' -o-aminoglycoside phosphotransferases were tolerant to Geneticin, G-418 Sulfate [2].
Treatment with 100 μg/mL Geneticin, G-418 Sulfate for 12 days did not lead to the death of all SSFs, but still 44.7% of SSFs survived. Continuous treatment with 200 μg/mL Geneticin, G-418 Sulfate for 12 days can lead to the death of all SSFs. Therefore, the minimum lethal concentration of Geneticin, G-418 Sulfate leading to the death of SSFs is 200 μg/mL[5]. Geneticin, G-418 Sulfate has antiviral activity against bovine viral diarrhea virus (BVDV). Geneticin, G-418 Sulfate can prevent cytopaplasia effect (CPE) caused by DENV-2 infection of BHK cells in a dose-dependent manner, with an EC50 value of 3μg/ml[3].
In vivo, Geneticin, G-418 Sulfate destabilizes mRNAs broadly, in that the majority of mRNAs in mESCs have reduced stability when mESCs are treated with Geneticin, G-418 Sulfate. The mRNAs with half-lives that are most reduced by treatment with Geneticin, G-418 Sulfate are enriched for select optimal codons, containing G/C at the wobble position[7].The expression of antioxidant stress kinase-related genes and apoptotic genes in donor cells treated with different concentrations of Geneticin, G-418 Sulfate significantly changed, but the DNA methylation level did not change.The in vitro development efficiency of nuclear transfer embryos from Geneticin, G-418 Sulfate-treated donor cells was significantly lower than that of controls[6].
Geneticin，G-418硫酸盐是一类氨基糖苷类抗生素，其结构与新霉素卡那霉素相似。它可以干扰真核细胞中的80S核糖体功能，从而阻止蛋白质合成并导致真核细胞死亡[1,4]。通常使用浓度为1至300μg/ml的Geneticin，G-418硫酸盐进行原核和真核克隆选择。临床分离的细菌株抗生素耐药性试验发现所有携带3'-O-氨基糖苷磷酸转移酶的菌株都对Geneticin，G-418硫酸盐具有耐受性[2]。
用100μg/mL的Geneticin，G-418硫酸盐处理12天并没有导致所有SSFs死亡，但仍有44.7%的SSFs存活。连续使用200μg/mL的Geneticin，G-418硫酸盐处理12天可以导致所有SSFs死亡。因此，导致SSFs死亡的最小致命浓度为200μg/mL[5]。 Geneticin，G-418硫酸盐对牛病毒性腹泻病毒（BVDV）具有抗病毒活性。 Geneticin，G-418硫酸盐可以以剂量依赖方式防止DENV-2感染BHK细胞引起的细胞质变性效应（CPE），EC50值为3μg/ml[3]。
在体内，Geneticin、G-418硫酸盐广泛破坏mRNA的稳定性，当mESCs接受Geneticin、G-418硫酸盐处理时，大多数mRNAs的稳定性都会降低。那些半衰期最受Geneticin、G-418硫酸盐处理影响的mRNAs富含选择性优化密码子，在wobble位点包含G/C[7]。经过不同浓度Geneticin、G-418硫酸盐处理后，供体细胞中与抗氧化应激激酶相关基因和凋亡基因表达显著改变，但DNA甲基化水平没有变化。从接受了Geneticin、G-418硫酸盐处理的供体细胞克隆出来的核移植胚胎在体外发育效率明显低于对照组[6]。
参考文献:
[1]: Bar-Nun S, Shneyour Y, et,al. G-418, an elongation inhibitor of 80 S ribosomes. Biochim Biophys Acta. 1983 Oct 13;741(1):123-7. doi: 10.1016/0167-4781(83)90018-0. PMID: 6193810.
[2]: Davies J, Jimenez A. A new selective agent for eukaryotic cloning vectors. Am J Trop Med Hyg. 1980 Sep;29(5 Suppl):1089-92. doi: 10.4269/ajtmh.1980.29.1089. PMID: 7001938.
[3]: Zhang XG, Mason PW, et,al. Antiviral activity of geneticin against dengue virus. Antiviral Res. 2009 Jul;83(1):21-7. doi: 10.1016/j.antiviral.2009.02.204. Epub 2009 Mar 11. PMID: 19501253; PMCID: PMC2694137.
[4]: Chen B., Shi X.Y., et,al. 2012, Cytoplasm vacuolization of fibroblasts during purification of Schwann cells by geneticin (G418): An optical microscope observation and analysis, Zhongguo Zhuzhi Gongcheng Yanjiu (Journal of Clinical Rehabilitative Tissue Engineering Research), 16 (14): 2593-2596
[5]: Wu Xian, JING Qian-ge, et,al. Study on the resistance of sheep skin fibroblasts to G418 and Blasticidin S [J].Genomics and applied biology,2019,38(03):1006-1011.
[6]: Li Jia-qi, Li Zi-Cong, et,al. Effects of G418 treatment on development efficiency of porcine cloned embryos in vitro [J]. Journal of south China agricultural university,2016,37(05):13-18.
[7]: Durmaz YT, Shatadal A, et,al. Geneticin reduces mRNA stability. PLoS One. 2022 Jul 28;17(7):e0272058. doi: 10.1371/journal.pone.0272058. PMID: 35901009; PMCID: PMC9333311.
| Cell experiment [1]: | |
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Cell lines |
Sheep skin fibroblasts, SSFs |
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Preparation Method |
The SSFs cultured in vitro were treated with different concentrations of single antibiotic Geneticin, G-418 Sulfate. After 12 days of treatment, the SSFs were digested, and the number of dead and alive cells was counted after placental blue staining to determine the lowest lethal dose of Geneticin, G-418 Sulfate leading to the death of all SSFs. |
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Reaction Conditions |
100-200ug/ml Geneticin, G-418 Sulfate for 1 day |
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Applications |
Treatment with 100 μg/mL Geneticin, G-418 Sulfate for 12 days did not lead to the death of all SSFs, but still (44.7 0.05)% of SSFs survived.Continuous treatment with 200 μg/mL Geneticin, G-418 Sulfate for 12 days can lead to the death of all SSFs. Therefore, the minimum lethal concentration of Geneticin, G-418 Sulfate leading to the death of SSFs is 200 μg/mL. |
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参考文献: [1]: Wu Xian, JING Qian-ge, et,al.Study on the resistance of sheep skin fibroblasts to G418 and Blasticidin S [J].Genomics and applied biology,2019,38(03):1006-1011. |
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