Molecular Biology

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    Protocols in Current Issue
    Enterovirus Competition Assay to Assess Replication Fitness
    Authors:  Valeria Lulla and Andrew E. Firth, date: 05/20/2019, view: 76, Q&A: 0
    [Abstract] In virology the difference between the fitness of two viruses can be determined by using various methods, such as virus titer, growth curve analysis, measurement of virus infectivity, analysis of produced RNA copies and viral protein production. However, for closely performing viruses, it is often very hard to distinguish the differences. In vitro ...
    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: 424, 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 ...
    Identification of RNase-sensitive LINE-1 Ribonucleoprotein Interactions by Differential Affinity Immobilization
    Authors:  Hua Jiang, Martin S. Taylor, Kelly R. Molloy, Ilya Altukhov and John LaCava, date: 04/05/2019, view: 609, Q&A: 0
    [Abstract] Long Interspersed Nuclear Element-1 (LINE-1, L1) constitutes a family of autonomous, self-replicating genetic elements known as retrotransposons. Although most are inactive, copious L1 sequences populate the human genome. L1s proliferate in a ‘copy-and-paste’ fashion through an RNA intermediate; a full-length L1 transcript is ~6,000 nucleotides ...
    Preparation of RNA 3’ End Sequencing Libraries of Total and 4-thiouracil Labeled RNA for Simultaneous Measurement of Transcription, RNA Synthesis and Decay in S. cerevisiae
    Authors:  Manfred Schmid, Agnieszka Tudek and Torben Heick Jensen, date: 03/20/2019, view: 548, Q&A: 0
    [Abstract] Cellular RNA levels are determined by the rates of RNA transcription from the gene template and subsequent RNA stability. Knowledge about both transcription and RNA decay is, therefore, necessary to interpret RNA levels and gene expression, especially during cellular processes where these parameters change. Numerous experimental strategies have ...
    Detection of mRNA by Whole Mount in situ Hybridization and DNA Extraction for Genotyping of Zebrafish Embryos
    Authors:  Rachna Narayanan and Andrew C. Oates, date: 03/20/2019, view: 1120, Q&A: 0
    [Abstract] In situ hybridization is used to visualize the spatial distribution of gene transcripts in tissues and in embryos, providing important information about disease and development. Current methods involve the use of complementary riboprobes incorporating non-radioactive labels that can be detected by immunohistochemistry and coupled to ...
    Quantification of Queuosine Modification Levels in tRNA from Human Cells Using APB Gel and Northern Blot
    Authors:  Zaneta Matuszek and Tao Pan, date: 03/20/2019, view: 491, Q&A: 0
    [Abstract] Queuosine (Q) is a hypermodified base in the wobble anticodon position of tRNAs coding for the amino acids Tyr, His, Asn, and Asp. tRNA Q-modification is introduced by a queuine tRNA-ribosyltransferase (TGT) that replaces the guanine base at G34 at these tRNAs with the modified base. tRNA Q-modification is widely distributed among prokaryotic and ...
    QsRNA-seq: A protocol for Generating Libraries for High-throughput Sequencing of Small RNAs
    Authors:  Alla Fishman and Ayelet T. Lamm, date: 03/05/2019, view: 742, Q&A: 0
    [Abstract] Small RNAs (sRNAs) are 20-30 nt long non-coding RNA molecules that regulate essentially all cellular processes. Besides being an intensively studied topic in academic research, sRNAs also hold a promise as clinical biomarkers. While the need for expressional profiling of sRNAs is growing, preparation of sRNA libraries for high-throughput ...
    pNP Transgenic RNAi System Manual in Drosophila
    Authors:  Fang Wang, Huan-Huan Qiao, Rong-Gang Xu, Jin Sun, Ruibao Zhu, Decai Mao and Jian-Quan Ni , date: 02/05/2019, view: 684, Q&A: 0
    [Abstract] Much of our knowledge about the mechanisms underlying biological processes relies on genetic approaches, whereby gene activity is reduced and the phenotypic consequences of perturbation are analyzed in detail. For functional genomic studies, a specific, systematic, and cost-effective manner is critical. Transgenic RNAi system is the top priority ...
    Gene Mapping by RNA-sequencing: A Direct Way to Characterize Genes and Gene Expression through Targeted Queries of Large Public Databases
    Author:  Peter Rotwein, date: 01/05/2019, view: 960, Q&A: 0
    [Abstract] Recent advances in genomics present new opportunities for enhancing knowledge about gene regulation and function across a wide spectrum of organisms and species. Understanding and evaluating this information at the individual gene level is challenging, and not only requires extracting, collating and interpreting data from public genetic ...
    Variable Dose Analysis: A Novel High-throughput RNAi Screening Method for Drosophila Cells
    Authors:  Katarzyna Sierzputowska, Chris R. Baxter and Benjamin E. Housden, date: 12/20/2018, view: 874, Q&A: 0
    [Abstract] Genetic screens are a powerful approach to identify previously uncharacterized genes involved in specific biological processes. Several technologies have been developed for high-throughput screens using reagents such as RNAi or CRISPR, and each approach is associated with specific advantages and disadvantages. Variable Dose Analysis (VDA), is an ...



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