Rapid and Specific Purification of RNA by PolyUracil Membranes

Year 2020, Volume: 10 Issue: 2, 466 - 482, 30.12.2020

Canan Armutcu

https://doi.org/10.37094/adyujsci.780049

Abstract

Ribonucleic acid (RNA) plays a critically important role in cellular defense, deoxyribonucleic acid (DNA) replication, transcription, and gene expression for living organisms as well as, a lot of diseases such as cancer, immunodeficiency, tumors has been associated with RNA’s disruption. For this purpose, supermacropores membranes were designed to purify RNA using nucleotide-based ligand. In this study, polymerizable uracil monomer as uracil methacrylate (UraM) was synthesized, and 2-hydroxyethyl methacrylate (HEMA)-based membranes [poly(HEMA-UraM)] were prepared by bulk polymerization under partially frozen conditions by copolymerization of monomers, UraM, and HEMA. These membranes were characterized via swelling studies, Fourier transform infrared spectroscopy (FTIR), and scanning electron microscopy (SEM). To optimize separation conditions, effects of pH, initial RNA concentration, time, and temperature on RNA adsorption capacity were examined. Maximum adsorption of RNA on poly(HEMA-UraM) membrane was found to be 15.52 mg/g for 0.5 mg/mL RNA initial concentration at 25.0°C with an optimum pH of 7.0. After ten repetitive adsorption-desorption cycles, the RNA adsorption capacity decreased only 3.68%.

Keywords

Uracil methacrylate, Ribonucleic acid, Membrane, Nucleotide, Purification

Thanks

C. Armutcu thanks Sena Piskin, Mirac Tuysuz and Dr. E. Ozgur for their valuable help during the study.

PoliUrasil Membranlar ile Hızlı ve Spesifik RNA Saflaştırılması

Abstract

Ribonükleik asit (RNA), hücresel savunma, deoksiribonükleik asit (DNA) replikasyonu, transkripsiyon, canlı organizmalar için gen ekspresyonununda önemli bir rol oynar, bunun yanı sıra kanser, immün yetmezlik, tümör gibi birçok hastalık RNA'nın bozulmasıyla ilişkililendirilmiştir. Bu amaçla, RNA saflaştırmak için nükleotid bazlı ligand kullanılarak gözenekli membranlar tasarlanmıştır. Bu çalışmada, polimerize edilebilir urasil monomeri urasil metakrilat (UraM) olarak sentezlenmiş, 2-hidroksietil metakrilat (HEMA) bazlı membranlar [poli(HEMA-UraM)] UraM ve HEMA monomerlerinin kopolimerizasyonu ile kısmen dondurulmuş koşullar altında yığın polimerizasyonu ile hazırlanmıştır. Bu membranlar şişme çalışması, Fourier dönüşümlü kızılötesi spektroskopisi (FTIR) ve taramalı elektron mikroskobu (SEM) ile karakterize edilmiştir. Ayırma koşullarını optimize etmek için, pH, başlangıç RNA konsantrasyonu, süre ve sıcaklığın RNA adsorpsiyon kapasitesi üzerindeki etkileri incelenmiştir. Poli(HEMA-UraM) membranın maksimum RNA adsorpsiyonu 25.0 °C, pH 7.0, 0.5 mg/mL RNA başlangıç derişiminde 15.52 mg/g olduğu bulunmuştur. On tekrarlı adsorpsiyon-desorpsiyon döngüsünden sonra, RNA adsorpsiyon kapasitesi yalnızca %3.68 oranında azaldığı gözlenmiştir.Membran;

Keywords

Membran, Urasil metakrilat, Nükleotid, Ribonükleik asit, Saflaştırma

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