Synthesis, deprotection, analysis and purification of RNA and ribosomes

作者：Francine E. Wincott, Anthony B. DiRenzo, Chris S. Shaffer, Susan Grimm, Danuta Tracz, Christopher Thomas Workman, David Sweedler, Carolyn Gonzalez, Stephen A. Scaringe, Nassim Usman · 发表于：Nucleic Acids Research · 年份：1995 · DOI：10.1093/nar/23.14.2677 · 被引用次数：398 · 研究领域：DNA and Nucleic Acid Chemistry、RNA and protein synthesis mechanisms、Advanced biosensing and bioanalysis techniques

Improvements in the synthesis, deprotection and purification of oligoribonucleotides are described. These advances allow for reduced synthesis and deprotection times, while improving product yield. Coupling times are reduced by half using 5-ethylthio-1H-tetrazole (S-ethyltetrazole) as the activator. Base and 2'-O-t-butyldimethylsilyl deprotection with methylamine (MA) and anhydrous triethylamine/hydrogen fluoride in N-methylpyrrolidinone (TEA.HF/NMP), respectively, requires a fraction of the time necessitated by current standard methods. In addition, the ease of oligoribonucleotide purification and analysis have been significantly enhanced using anion exchange chromatography. These new methods improve the yield and quality of the oligoribonucleotides synthesized. Hammerhead ribozymes synthesized utilizing the described methods exhibited no diminution in catalytic activity.