Open AccessSynthesis, deprotection, analysis and purification of RNA and ribosomes
Author(s) -
Francine E. Wincott,
Anthony B. DiRenzo,
Chris S. Shaffer,
Susan Grimm,
Danuta Tracz,
Christopher T. Workman,
David Sweedler,
C. Gonzalez,
Stephen A. Scaringe,
Nassim Usman
Publication year - 1995
Publication title -
nucleic acids research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 9.008
H-Index - 537
eISSN - 1362-4954
pISSN - 0305-1048
DOI - 10.1093/nar/23.14.2677
Subject(s) - triethylamine , biology , yield (engineering) , methylamine , combinatorial chemistry , ribosome , rna , ribozyme , chromatography , biochemistry , organic chemistry , chemistry , materials science , gene , metallurgy
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.