Method for preparing deoxyribonucleotide
A deoxynucleotide and reaction system technology, applied in the field of deoxynucleotide preparation, can solve the problems of uneasy control of reaction conditions, low substrate conversion rate, low product yield, etc., and achieve a wide range of system pH and temperature, High activity, wide range of pH and temperature adaptability
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Embodiment 1
[0013] The reaction system adopts acetic acid buffer system, pH 4.0. Add to 200mM acetate buffer: hydrolyze the substrate deoxyribonucleic acid to a final concentration of 1mg / mL, Cu 2+ The final concentration of glucosamine oligosaccharide is 1 μM, and the final concentration of glucosamine oligosaccharide is 0.1 μM, so that the final reaction volume is 5 mL, the molecular weight of glucosamine oligosaccharide in the reaction system is 340-984 Da, and the reaction temperature is 40°C. React for 12 hours, use deionized water to pass the reaction solution through an anion chromatography column with a diameter of 2.5×25cm for adsorption, the flow rate is 2mL / Min, balance with deionized water to remove unadsorbed impurities, and then use 1.8mM HCl deionized aqueous solution The dCMP was eluted, the dAMP was eluted with 2.8 mM HCl in deionized water, the dTMP was eluted with 36 mM NaCl in deionized water, and the dGMP was eluted with 5 mM HCl, 20 mM NaCl in deionized water. Final...
Embodiment 2
[0015] The reaction system adopts acetic acid buffer system, pH6.0. Add to 100mM acetate buffer: hydrolyze the substrate deoxyribonucleic acid to a final concentration of 30mg / mL, Cu 2+ The final concentration of glucosamine oligosaccharide is 15 μM, and the final concentration of glucosamine oligosaccharide is 5 μM, so that the final reaction volume is 5 mL, the molecular weight of glucosamine oligosaccharide in the reaction system is 340-984 Da, and the reaction temperature is 30° C. React for 4 hours, use deionized water to pass the reaction solution through an anion chromatography column with a diameter of 2.5×25cm for adsorption, the flow rate is 2mL / Min, balance with deionized water to remove unadsorbed impurities, and then use 1.8mM HCl deionized aqueous solution The dCMP was eluted, the dAMP was eluted with 2.8 mM HCl in deionized water, the dTMP was eluted with 36 mM NaCl in deionized water, and the dGMP was eluted with 5 mM HCl, 20 mM NaCl in deionized water. Finall...
Embodiment 3
[0017] The reaction system adopts Tris-HCl buffer system, pH7.5. Add to 200mM Tris-HCl buffer: hydrolyze the substrate deoxyribonucleic acid to a final concentration of 100mg / mL, Fe 2+ The final concentration of glucosamine oligosaccharide is 1 μM, the final reaction volume is 5 mL, the molecular weight of glucosamine oligosaccharide in the reaction system is 340-984 Da, and the reaction temperature is 20°C. After reacting for 6 hours, dilute the reaction solution 100 times with deionized water and then pass it through an anion chromatography column with a diameter of 2.5×25cm for adsorption at a flow rate of 2mL / Min, balance with deionized water to remove unadsorbed impurities, and then use 1.8mM The dCMP was eluted with deionized HCl, the dAMP was eluted with 2.8 mM HCl deionized water, the dTMP was eluted with 36 mM NaCl deionized water, and the dGMP was eluted with 5 mM HCl, 20 mM NaCl deionized water. Finally, the ion exchange medium is regenerated with 1M hydrochloric a...
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