Preparation of modified long-mer RNAs and analysis of FMN binding to the<i>ypaA</i>aptamer from<i>B. subtilis</i>

Frommer, Jennifer; Hieronymus, Robert; Arunachalam, Tamil Selvi; Heeren, Sabine; Jenckel, Maria; Strahl, Anne; Appel, Bettina; Müller, Sabine

doi:10.4161/rna.28526

Cited by 12 publications

(30 citation statements)

References 52 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Pre-orientation of 5′-p and 3′-OH of the RNA substrate is achieved ( a ) by hybridization to a splint. This strategy can be used for ligation with T4 DNA ligase (DNA splint required), T4 RNA ligase 1 (RNA splint required) and T4 RNA ligase 2 (DNA or RNA splint) ( 68 ); ( b ) by hairpin and linear helper oligonucleotides ( 87 ); ( c ) by a gap splint that upon hybridization leaves the terminal two or three nucleotides of both ends single stranded at the ligation junction ( 88 ); ( d ) by favorable intrinsic structures (i.e. dumbbell folds ( 17 )).…”

Section: Circularization In Vitromentioning

confidence: 99%

“…In addition to helper oligonucleotides that assist in ligation by interaction with the substrate in a distance from the ligation site (Figure 5b ), also splints may be used that bring the reactive ends close together, but leave the terminal two to three nucleotides of both donor and acceptor single stranded. This is a common procedure for ssRNA ligation with T4 RNA ligase 1 ( 88 , 89 ), and should be a suitable strategy also for circularization (Figure 5c ). Another scenario uses internal base pairing of the linear precursor to promote ligation (Figure 5d ).…”

Section: Circularization In Vitromentioning

confidence: 99%

See 1 more Smart Citation

RNA circularization strategies in vivo and in vitro

Petkovic

Müller

2015

Nucleic Acids Research

Self Cite

278

218

View full text Add to dashboard Cite

In the plenitude of naturally occurring RNAs, circular RNAs (circRNAs) and their biological role were underestimated for years. However, circRNAs are ubiquitous in all domains of life, including eukaryotes, archaea, bacteria and viruses, where they can fulfill diverse biological functions. Some of those functions, as for example playing a role in the life cycle of viral and viroid genomes or in the maturation of tRNA genes, have been elucidated; other putative functions still remain elusive. Due to the resistance to exonucleases, circRNAs are promising tools for in vivo application as aptamers, trans-cleaving ribozymes or siRNAs. How are circRNAs generated in vivo and what approaches do exist to produce ring-shaped RNAs in vitro? In this review we illustrate the occurrence and mechanisms of RNA circularization in vivo, survey methods for the generation of circRNA in vitro and provide appropriate protocols.

show abstract

Section: Circularization In Vitromentioning

confidence: 99%

Section: Circularization In Vitromentioning

confidence: 99%

RNA circularization strategies in vivo and in vitro

Petkovic

Müller

2015

Nucleic Acids Research

Self Cite

278

218

View full text Add to dashboard Cite

show abstract

“…For example, recently we have prepared a modified 129mer RNA for fluorescence studies by enzymatic and chemical ligation of 2 RNA fragments. 23 As an alternative to chemical synthesis, RNA can be produced by in vitro transcription, which allows preparation of RNAs of virtually unlimited length, however, requires post-synthetic functionalization strategies.…”

Section: Chain Assemblymentioning

confidence: 99%

“…23 A characteristic feature of in vitro transcribed RNA is the 5 0 -terminal triphosphate, because 5 0 -triphosphates of nucleosides are used as building blocks for enzymatic polymerization. As mentioned above, enzymatic end-joining for circularization mostly requires a 5 0 -monophosphate, and also chemical procedures rely on specific functionalities.…”

Section: Chain Assemblymentioning

confidence: 99%

“…Another option is the reaction of the unique primary alcohol at the 5 0 -end with triphenylphosphonium iodide to generate a good leaving group and thus making the 5 0 -terminus suitable for reaction with a nucleophile, as for example an azide. 23,24 The azide may be directly used for Click cyclization (see below) or, upon reduction to an amino group, for conjugation with a desired functionality via peptide chemistry. 25 Specific 5 0 -RNA functionalization can be also achieved by transcription priming, adding to the transcription mixture an initiator nucleotide that carries no triphosphate for polymerization and therefore can be used only for priming RNA synthesis at the 5 0 -end.…”

Section: Chain Assemblymentioning

confidence: 99%

See 1 more Smart Citation

In vitro circularization of RNA

Müller

Appel

2016

RNA Biology

Self Cite

View full text Add to dashboard Cite

Over the past 2 decades, different types of circular RNAs have been discovered in all kingdoms of life, and apparently, those circular species are more abundant than previously thought. Apart from circRNAs in viroids and viruses, circular transcripts have been discovered in rodents more than 20 y ago and recently have been reported to be abundant in many organisms including humans. Their exact function remains still unknown, although one may expect extensive functional studies to follow the currently dominant research into identification and discovery of circRNA by sophisticated sequencing techniques and bioinformatics. Functional studies require models and as such methods for preparation of circRNA in vitro. Here, we will review current protocols for RNA circularization and discuss future prospects in the field.Abbreviations: AMP, adenosine monophosphate; AppRNA, 5 0 -adenylated RNA; ATP, adenosine triphosphate; bp, base pair; CEV-A, citrus exocortis viroid strain A; circRNA, circular RNA; ciRNA, circular intronic RNA; CuAAC, cupper catalyzed azide alkyne cycloaddition; EDC, ethyl-3-(3 0 -dimethylaminopropyl)-carbodiimide; HIV, human immunodeficiency virus; IEciRNA, intron-exon circular RNA; nt, nucleotide; PIE, permuted intron exon; T4 Dnl, T4 DNA Ligase; T4 Rnl, T4 RNA Ligase

show abstract

Current Approaches for RNA Labeling in Vitro and in Cells Based on Click Reactions

Schulz

Rentmeister

2014

ChemBioChem

View full text Add to dashboard Cite

Over recent years, click reactions have become recognized as valuable and flexible tools to label biomacromolecules such as proteins, nucleic acids, and glycans. Some of the developed strategies can be performed not only in aqueous solution but also in the presence of cellular components, as well as on (or even in) living cells. These labeling strategies require the initial, specific modification of the target molecule with a small, reactive moiety. In the second step, a click reaction is used to covalently couple a reporter molecule to the biomolecule. Depending on the type of reporter, labeling by the click reaction can be used in many different applications, ranging from isolation to functional studies of biomacromolecules. In this minireview, we focus on labeling strategies for RNA that rely on the click reaction. We first highlight click reactions that have been used successfully to label modified RNA, and then describe different strategies to introduce the required reactive groups into target RNA. The benefits and potential limitations of the strategies are critically discussed with regard to possible future developments.

show abstract

Preparation of modified long-mer RNAs and analysis of FMN binding to theypaAaptamer fromB. subtilis

Cited by 12 publications

References 52 publications

RNA circularization strategies in vivo and in vitro

RNA circularization strategies in vivo and in vitro

In vitro circularization of RNA

Current Approaches for RNA Labeling in Vitro and in Cells Based on Click Reactions

Contact Info

Product

Resources

About