一种用C18柱纯化低浓度微量寡核苷酸的新方法

A New Method for Purifying Little Oligonucleotide from Low Concentration of Solution by C18 Column

通过比较4种不同的回收方法和C18硅胶4种不同的用量对回收的影响,提出了回收低浓度微量寡核苷酸的最佳方案:分别用1 mL乙腈、2 mL灭菌双蒸水和1 mL 10 mmol/L乙酸铵平衡装填10 mg硅胶的C18 Sep-Pak反相层析柱;再将寡核苷酸溶液通过C18柱;然后用1 mL 25 mmol/L碳酸氢铵洗脱C18柱,并以3000 r/m in离心2 m in;最后用1 mL 30%乙腈洗脱C18柱,将收集的溶液离心冻干后即得到纯化的寡核苷酸。该方法的回收率可达53.7%±3.0%。

By comparing 4 different recovery methods and four dosages of C18 silica gel, a best method was suggested for purifying little oligonucleotide from low concentration of solution. A C18 Sep - Pak cartridge filled with 10 mg silica gel was prepared by rinsing with 1 mL acetonitrile, 2 mL sterile ddH20 and 1 mL 10 mmol/L NH4AC subsequentially. The oligonucleotide solution was then applied to the cartridge. After that, the C18 column was washed by 1 mL 25mmol/L NH4HCO3. The column was then centrifugated at 3000 r/min for 2 rain. Finally, the oligonucleotide solution was eluted from the column by 1 mL 30% acetonitrile. The purified oligonucleotide was obtained through centrifugating and freezing the collected oligonucleotide solution. The recovery rate of oligonucleotide reached 53.7% ±3.0%.