ArticleRNA (New York, N.Y.)2016
Adenylylation of small RNA sequencing adapters using the TS2126 RNA ligase I.
Article in RNA (New York, N.Y.), 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 12 papers.
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Who cites it
12 citing papers in PubMed.
- High-resolution profiling of bacterial transcriptomes by SEnd-seq.The Biochemical journal · 2026Review
- Transcription attenuation amplifies collateral vulnerabilities in rifampicin-resistant Mycobacterium tuberculosis.Nature microbiology · 2026Article
- Temperature-Dependent tRNA Modifications in Bacillales.International journal of molecular sciences · 2024Article
- Characterisation and engineering of a thermophilic RNA ligase from Palaeococcus pacificus.Nucleic acids research · 2024Article
- Article
- Functional biology and biotechnology of thermophilic viruses.Essays in biochemistry · 2023Review
- Formamide significantly enhances the efficiency of chemical adenylation of RNA sequencing ligation adaptors.RNA (New York, N.Y.) · 2023Article
- Potential Applications of Thermophilic Bacteriophages in One Health.International journal of molecular sciences · 2023Review
- Direct adenylation from 5'-OH-terminated oligonucleotides by a fusion enzyme containing Pfu RNA ligase and T4 polynucleotide kinase.Nucleic acids research · 2022Article
- cyPhyRNA-seq: a genome-scale RNA-seq method to detect active self-cleaving ribozymes by capturing RNAs with 2',3'RNA biology · 2021Article
- Full-length RNA profiling reveals pervasive bidirectional transcription terminators in bacteria.Nature microbiology · 2019Article
- Small RNA-seq: The RNA 5'-end adapter ligation problem and how to circumvent it.Journal of biological methods · 2019Article
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2 authors.
Funding
Abstract
Many high-throughput small RNA next-generation sequencing protocols use 5' preadenylylated DNA oligonucleotide adapters during cDNA library preparation. Preadenylylation of the DNA adapter's 5' end frees from ATP-dependence the ligation of the adapter to RNA collections, thereby avoiding ATP-dependent side reactions. However, preadenylylation of the DNA adapters can be costly and difficult. The currently available method for chemical adenylylation of DNA adapters is inefficient and uses techniques not typically practiced in laboratories profiling cellular RNA expression. An alternative enzymatic method using a commercial RNA ligase was recently introduced, but this enzyme works best as a stoichiometric adenylylating reagent rather than a catalyst and can therefore prove costly when several variant adapters are needed or during scale-up or high-throughput adenylylation procedures. Here, we describe a simple, scalable, and highly efficient method for the 5' adenylylation of DNA oligonucleotides using the thermostable RNA ligase 1 from bacteriophage TS2126. Adapters with 3' blocking groups are adenylylated at >95% yield at catalytic enzyme-to-adapter ratios and need not be gel purified before ligation to RNA acceptors. Experimental conditions are also reported that enable DNA adapters with free 3' ends to be 5' adenylylated at >90% efficiency.
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