Extraction of urinary cell-free DNA by using triamine-modified silica particles for liquid biopsy
Extraction of urinary cell-free DNA by using triamine-modified silica particles for liquid biopsy
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DOI：
10.1007/s00216-020-02784-5
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发表时间：
2020-07-02
影响因子：
4.3
通讯作者：
Karasawa, Koji
中科院分区：
文献类型：
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作者：
Kato, Masaru;Fujita, Yumi;Karasawa, Koji
尿液中存在约200 bp的无细胞DNA（cfDNA）作为液体活检的生物标志物引起了人们的注意。然而，它目前仅可用于诊断其中大量cfDNA在尿液中排泄的癌症。因此，开发一种提取尿液中少量存在的cfDNA的有效方法对于诊断许多其他疾病至关重要。我们研究了粒径，小孔径（表面积），和多孔二氧化硅颗粒的表面改性对DNA提取效率的影响。我们的观察表明cfDNA可以被叔胺修饰的颗粒捕获，然后通过用碳酸氢钠（pH 11）反复洗涤从颗粒中除去。使用这种方法，使用30 mg三胺修饰的颗粒，我们成功地从15 mL尿液中提取了几百纳克的cfDNA。在15 mL尿样中可检测到约67 fg/mL的龋DNA（211 bp），表明该方法可用于基于cfDNA的液体活检的遗传标记物提取。
The presence of approximately 200-bp cell-free DNA (cfDNA) in the urine has attracted attention as a biomarker for liquid biopsy. However, it is currently useful only for diagnoses of cancers in which a large amount of cfDNA is excreted in the urine. Therefore, the development of an efficient method for extracting cfDNA existing in small amounts in the urine is essential for diagnosing many other diseases. We examined the effect of particle size, small pore size (surface area), and surface modification of porous silica particles on the efficiency of DNA extraction. Our observations suggested that cfDNA could be captured by tertiary amine-modified particles and then removed from the particles by repeatedly washing with sodium bicarbonate (pH 11). Using this method with 30 mg of triamine-modified particles, we succeeded in extracting a few hundred nanograms of cfDNA from 15 mL urine. Furthermore, we could detect similar to 67 fg/mL caries DNA (211 bp) in 15 mL urine sample, suggesting that this method may be suitable for the extraction of genetic biomarkers for cfDNA-based liquid biopsy.