Titlebook: Recombinant and In Vitro RNA Synthesis; Methods and Protocol Graeme L. Conn Book 2012 Springer Science+Business Media, LLC 2012 RNA purific

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,,-Acting Ribozymes for the Production of RNA In Vitro Transcripts with Defined 5′ and 3′ Ends, at both the 5′- and 3′-ends. This chapter describes the use of .-acting ribozymes, 5′-end hammerhead (HH) and 3′-end hepatitis delta virus (HDV), for direct transcriptional processing to yield target RNAs with precisely defined ends. The method is focused on the use of the pRZ and p2RZ plasmids tha

phase-2-enzyme

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Rapid Preparation of RNA Samples Using DNA-Affinity Chromatography and DNAzyme Methods,ription method results in heterogeneity at the RNA 3′-terminus. RNA purification requires single-nucleotide resolution to separate the transcript of the correct length from the aborted or add-on transcripts that are usually present in comparable amounts. Here, we describe an RNA preparation method t

Predigest

,Preparation of λN-GST Fusion Protein for Affinity Immobilization of RNA, Affinity purification is particularly advantageous because it can be performed in a few hours under non-denaturing conditions. However, the performance of affinity purification methods can vary tremendously depending on the RNA immobilization matrix. It was previously shown that RNA immobilization

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Engulf

In Vitro Transcription of Modified RNAs,o standard transcription protocols. RNAs containing a single phosphate or a cap structure at their 5′ ends can readily be generated either co-transcriptionally or through enzymatic treatments of transcription products. Likewise, a variety of modified bases, including fluorescent or biotinylated spec

medium

End-Labeling Oligonucleotides with Chemical Tags After Synthesis,le nucleic acid detection, yield information about its state, and can serve as a handle by which the nucleic acid and associated factors can be purified from a mixture. Radioactive phosphate is commonly added to the 5′ or 3′ end of an oligonucleotide post synthesis using enzyme-catalyzed reactions.

abduction

High-Purity Enzymatic Synthesis of Site-Specifically Modified tRNA,ndent on the presence of modified nucleotides in tRNA, each of which performs a distinct function. To better understand how individual modifications modulate tRNA function, a method to isolate and purify a site-specifically modified tRNA is essential. This chapter describes an enzymatic method to sy

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