[7] Joining of RNAs by splinted ligation
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A review on recent advances in methods for site-directed spin labeling of long RNAs
2023, International Journal of Biological MacromoleculesStructural basis of microRNA biogenesis by Dicer-1 and its partner protein Loqs-PB
2022, Molecular CellCitation Excerpt :RNAs for dicing assays (Table S2, Integrated DNA Technologies) were 5′ 32P radiolabeled using [γ-32P]ATP (6000 Ci/mmol) (PerkinElmer, Waltham, MA) and T4 polynucleotide kinase (NEB, Ipswich, MA), and then gel-purified. Substrates containing two 32P-radiolabeled phosphates were prepared by DNA-splinted ligation (Moore and Sharp, 1993; Moore and Query, 2000). Synthetic hairpins for substrate competition assays (Table S2) were ordered from Integrated DNA Technologies as HPLC-purified RNAs.
Circular RNA: An emerging frontier in RNA therapeutic targets, RNA therapeutics, and mRNA vaccines
2022, Journal of Controlled ReleaseCitation Excerpt :For example, DNA and RNA ligases originating from T4 bacteriophage are employed to ligate linear RNAs with terminal monophosphate. Due to the high selectivity of T4 DNA ligase, only RNAs that are entirely complementary to the corresponding DNA splints will be ligated [75]. T4 DNA ligase-based RNA circularization is particularly useful to precisely synthesize circRNA using precursor linear RNAs synthesized by IVT, in which the run-off properties of RNA polymerases often cause heterogeneity of RNA termini [76].
The design and synthesis of circular RNAs
2021, MethodsCitation Excerpt :This specificity is particularly desirable when the linear precursor RNA is synthesized through IVT, which results in considerable 3′ terminal heterogeneity [35,36]. Since the turnover efficiency of DNA ligase I on RNA substrates is low, a large amount of enzyme is often needed to achieve RNA ligation, particularly when compared to use of dedicated RNA ligases [47]. Compared to DNA ligase 1, T4 RNA ligase 1 works only on single-stranded substrates and exhibits lower reaction specificity [51].