真核细胞中基于自旋柱的RNA纯化:一篇教育研究文章

M. Abdelfattah, Mohamed R. Elnagar, M. Ibrahim, M. Albert, Maya Talal, Reham Helwa
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摘要

20年前,在世界范围内进行分子生物学研究是非常昂贵的。后来,在分配了许多试剂的专有技术之后,以合理的成本在该领域开展工作变得更加可行。此外,酶、试剂盒、实验室耗材和试剂的大规模生产优化使研究和诊断领域发生了重大变革。因此,在任何分子生物学实验室拥有负担得起的技术本身就是一个目标,特别是在发展中国家。新冠肺炎疫情后,我国对分子生物学试剂的需求激增。然而,耗材价格、订单延迟和短缺是疫情后的突出问题。因此,在本研究中,我们研究了从HepG2和huh7细胞中纯化RNA的自制溶液。用自制溶液成功纯化RNA,并用qRT-PCR扩增。然而,DNA污染可以通过DNA酶I酶切或在RT-PCR的外显子-外显子连接处设计合适的引物以及精确的归一化策略来消除。
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Composing solutions for spin-column based RNA purification from eukaryotic cells: An educational research article
Twenty years ago, working with molecular biology research was very costly worldwide. Later on, after distributing the know-how of many reagents, it becomes more feasible to work in this field with reasonable costs. In addition, the optimization of large-scale production of enzymes, kits, lab consumables, and reagents made a big revolution in research and diagnostics. So, having affordable technologies in any molecular biology laboratory is an aim itself, especially in developing countries. After Covid-19 crisis, the need of molecular biology reagents is in surge. However, consumables’ prices, delayed orders, and shortage are the prominent issues after the pandemic. Thus, in the present study, homemade solutions were investigated for RNA purification from HepG2 and huh7 cells. The RNA was successfully purified by the homemade solutions and amplified using qRT-PCR. However DNA contamination was encountered which could be eliminated simply by DNase I digestion or designing proper primers in exon-exon junction for RT-PCR, in addition to precise normalization strategy.
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