Biochemistry

Categories

    Protocols in Current Issue
    Northern Blot with IR Fluorescent Probes: Strategies for Probe Preparation
    Authors:  Christopher Fields, Peike Sheng, Bret Miller, Tianqi Wei and Mingyi Xie, date: 04/20/2019, view: 456, Q&A: 0
    [Abstract] Northern blot is a molecular biology technique that can detect, quantify, and determine the molecular weight of RNA. Recently, we published a protocol utilizing near-infrared (IR) fluorescent probes in Northern blot (irNorthern). Our method is as sensitive as other non-radioactive methods but is more straightforward and versatile. Additionally, we ...
    Preparation of Sequencing RNA Libraries through Chemical Cross-linking Coupled to Affinity Purification (cCLAP) in Saccharomyces cerevisiae
    Authors:  Congwei Wang, Julie Weidner and Anne Spang, date: 10/05/2018, view: 1679, Q&A: 0
    [Abstract] Ribonucleoprotein particles (mRNPs) are complexes consisting of mRNAs and RNA-binding proteins (RBPs) which control mRNA transcription localization, turnover, and translation. Some mRNAs within the mRNPs have been shown to undergo degradation or storage. Those transcripts can lack general mRNA elements, like the poly(A) tail or 5’ cap structure, ...
    Enzymatic Synthesis and Fractionation of Fluorescent PolyU RNAs
    [Abstract] The physical properties of viral-length polyuridine (PolyU) RNAs, which cannot base-pair and form secondary structures, are compared with those of normal-composition RNAs, composed of comparable numbers of each of A, U, G and C nucleobases. In this protocol, we describe how to synthesize fluorescent polyU RNAs using the enzyme polynucleotide ...
    Purification of RNA Mango Tagged Native RNA-protein Complexes from Cellular Extracts Using TO1-Desthiobiotin Fluorophore Ligand
    Authors:  Shanker Shyam Sundhar Panchapakesan, Sunny C. Y. Jeng and Peter J. Unrau, date: 04/05/2018, view: 2004, Q&A: 0
    [Abstract] A native purification strategy using RNA Mango for RNA based purification of RNA-protein complexes is described. The RNA Mango aptamer is first genetically engineered into the RNA of interest. RNA Mango containing complexes obtained from cleared cellular native extracts are then immobilized onto TO1-Desthiobiotin saturated streptavidin agarose ...
    Accurate, Streamlined Analysis of mRNA Translation by Sucrose Gradient Fractionation
    Authors:  Soufiane Aboulhouda, Rachael Di Santo, Gabriel Therizols and David Weinberg, date: 10/05/2017, view: 4707, Q&A: 0
    [Abstract] The efficiency with which proteins are produced from mRNA molecules can vary widely across transcripts, cell types, and cellular states. Methods that accurately assay the translational efficiency of mRNAs are critical to gaining a mechanistic understanding of post-transcriptional gene regulation. One way to measure translational efficiency is to ...
    Mapping RNA Sequences that Contact Viral Capsid Proteins in Virions
    Authors:  C. Cheng Kao, Ella Chuang, James Ford, Jie Huang, Ram Podicheti and Doug B. Rusch, date: 07/20/2017, view: 3651, Q&A: 0
    [Abstract] We have adapted the methodology of CLIP-seq (Crosslinking-Immunoprecipitation and DNA Sequencing) to map the segments of encapsidated RNAs that contact the protein shells of virions. Results from the protocol report on the RNA sequences that contact the viral capsid.
    Biotinylated Micro-RNA Pull Down Assay for Identifying miRNA Targets
    Authors:  Pornima Phatak and James M Donahue, date: 05/05/2017, view: 7360, Q&A: 1
    [Abstract] microRNA (miRNA) directly associates with its target transcripts (mRNA). This protocol describes a method for detection of direct interaction between miRNA and mRNA. The result of interaction helps screening the specific target mRNAs for a miRNA.
    RNA-dependent RNA Polymerase Assay for Hepatitis E Virus
    Authors:  Vidya P. Nair, Saumya Anang, Akriti Srivastava and Milan Surjit, date: 04/05/2017, view: 4551, Q&A: 0
    [Abstract] RNA-dependent RNA polymerase (RdRp) is essential for the replication of viral RNA for RNA viruses. It synthesizes the complementary strand of viral genomic RNA, which is used subsequently as a template to generate more copies of viral genome. This assay measures activity of the hepatitis E virus (HEV) RdRp. In contrast to protocols available to ...
    RNA Strand Displacement Assay for Hepatitis E Virus Helicase
    Authors:  Vidya P. Nair and Milan Surjit, date: 04/05/2017, view: 4222, Q&A: 0
    [Abstract] The hepatitis E virus (HEV) helicase uses ATP to unwind the RNA duplexes. This is an essential step for viral replication. This protocol aims to measure the double strand RNA unwinding activity of the HEV helicase.
    Polysome Analysis
    Authors:  Dipak Kumar Poria and Partho Sarothi Ray, date: 03/20/2017, view: 6710, Q&A: 1
    [Abstract] Polysome analysis is a method to separate mRNAs from a cell into actively translating and non-translating fractions depending on their association with polysomes. By this protocol, cell lysates are fractionated by sucrose density gradient ultracentrifugation. Free mRNA fraction and various ribosomal fractions, such as 40S, 60S, monosomes and ...



    We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.