Equimolar addition of oligoribonucleotides with T4 RNA ligase
نویسندگان
چکیده
T4 induced RNA ligase will join equimolar concentrations of two oligoribonucleotides, (Ap)3C and p(Up) 5, to form a single product, (Ap)3Cp(Up) 5, in high yield. The presence of the 3' phosphate on p(Up)5 prevents the oligomer from adding to itself. The pH optimum of the reaction is about 7.5, but less of the undesirable adenylated intermediate, App(Up) 5, forms at pH 8.2. The reaction rate is a linear function of oligomer concentration from 3 micronM to 0.6 mM. The data suggest that T4 RNA ligase will be a useful enzyme for the synthesis of oligomers of defined sequence.
منابع مشابه
Addition of mononucleotides to oligoribonucleotide acceptors with T4 RNA ligase.
RNA ligase induced by bacteriophage T4 catalyzed the addition of nucleoside 5',3'-diphosphates to oligoribonucleotide acceptors in the presence of ATP. The reactions proceeded in equimolar concentrations of donors and acceptors. Several oligonucleotides of defined sequence were synthesized by this method in yields of 28-68%. The enzyme required the phosphate ester at the 3' position of the dono...
متن کاملInitiation of synthesis of messenger RNA of deoxynucleotide kinase by oligoribonucleotides.
The effects of nucleoside triphosphates and oligoribonucleotides on the initiation of synthesis of messenger RNA of the T4 phage-specific enzyme, deoxynucleotide kinase, have been studied. The procedure involved incubation of T4 DNA, purified RNA polymerase from Escherichia coli, and selected nucleotide compounds during a brief period to permit initiation of RNA synthesis. Further initiation wa...
متن کاملAn RNA ligase-mediated method for the efficient creation of large, synthetic RNAs.
RNA ligation has been a powerful tool for incorporation of cross-linkers and nonnatural nucleotides into internal positions of RNA molecules. The most widely used method for template-directed RNA ligation uses DNA ligase and a DNA splint. While this method has been used successfully for many years, it suffers from a number of drawbacks, principally, slow and inefficient product formation and sl...
متن کاملIsolation and characterization of an RNA ligase from HeLa cells.
An RNA ligase has been purified from HeLa cells, which catalyzes the intra- and intermolecular ligation of linear RNA substrates possessing 5'-hydroxyl and 2',3'-cyclic phosphate termini in the presence of ATP or dATP. In this reaction, the 2',3'-cyclic phosphate is incorporated into a 3'-5'-phosphodiester bond, in agreement with the findings of Filipowicz et al. [Filipowicz, W., Konarska, M., ...
متن کاملCovalent attachment of polyribonucleotides to polydeoxyribonucleotides catalyzed by deoxyribonucleic acid ligase.
DNA ligase isolated from Escherichia coli or phage T4-infected E. coli catalyzes the covalent joining of the 5’-phosphate termini of polydeoxyribonucleotides and 3’-hydroxyl termini of polyribonucleotides. This reaction occurs with poly(dA) and poly(A) in the presence of poly(dT). The 5’-phosphate terminus of poly[d(A-T)] can be linked by an intramolecular reaction to its 3’-hydroxyl terminus s...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
عنوان ژورنال:
- Nucleic acids research
دوره 4 1 شماره
صفحات -
تاریخ انتشار 1977