Molecular Biology

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    Protocols in Current Issue
    A Highly Sensitive Method to Efficiently Profile the Histone Modifications of FFPE Samples
    Authors:  Linxuan Zhao, Vamsi Krishna Polavarapu, Ram Prakash Yadav, Pengwei Xing and Xingqi Chen, date: 05/20/2022, view: 480, Q&A: 0
    [Abstract]

    The majority of biopsies in both basic research and translational cancer studies are preserved in the format of archived formalin-fixed paraffin-embedded (FFPE) samples. Profiling histone modifications in archived FFPE tissues is critically important to understand gene regulation in human disease. The required input for current genome-wide

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    A Molecular Cloning and Sanger Sequencing-based Protocol for Detecting Site-specific DNA Methylation
    Authors:  Wei Guo, Anthony Cannon and Damon Lisch, date: 05/05/2022, view: 519, Q&A: 0
    [Abstract]

    DNA methylation is a conserved chemical modification, by which methyl groups are added to the cytosine of DNA molecules. Methylation can influence gene expression without changing the sequence of a particular gene. This epigenetic effect is an intriguing phenomenon that has puzzled biologists for years. By probing the temporal and spatial patterns

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    An Improved EMSA-based Method to Prioritize Candidate cis-REs for Further Functional Validation
    Authors:  Ting Wu and Gang Li, date: 04/20/2022, view: 464, Q&A: 0
    [Abstract]

    Cells are the complex product of gene expression programs that involve the coordinated transcription of thousands of genes controlled by cis-regulatory elements (cis-REs). Therefore, identification of cis-REs is the key to decipher the mechanisms underlying the regulation of gene expression. Here, we describe a simple and time-effective protocol

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    An Optimized Tat/Rev Induced Limiting Dilution Assay for the Characterization of HIV-1 Latent Reservoirs
    [Abstract]

    The administration of antiretroviral therapy (ART) leads to a rapid reduction in plasma viral load in HIV-1 seropositive subjects. However, when ART is suspended, the virus rebounds due to the presence of a latent viral reservoir. Several techniques have been developed to characterize this latent viral reservoir. Of the various assay formats

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    Conditional Human BRD4 Knock-In Transgenic Mouse Genotyping and Protein Isoform Detection
    Authors:  Michael Paul Lewis, Shwu-Yuan Wu and Cheng-Ming Chiang, date: 04/05/2022, view: 451, Q&A: 0
    [Abstract]

    Bromodomain-containing protein 4 (BRD4) is an acetyl-lysine reader protein and transcriptional regulator implicated in chromatin dynamics and cancer development. Several BRD4 isoforms have been detected in humans with the long isoform (BRD4-L, aa 1-1,362) playing a tumor-suppressive role and a major short isoform (BRD4-S, aa 1-722) having

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    A Rapid FRET Real-Time PCR Protocol for Simultaneous Quantitative Detection and Discrimination of Human Plasmodium Parasites
    [Abstract]

    Malaria is the most important parasitic disease worldwide, and accurate diagnosis and treatment without delay are essential for reducing morbidity and mortality, especially in P. falciparum malaria. Real-time PCR is highly sensitive and highly specific, therefore an excellent diagnostic tool for laboratory detection and species-specific

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    Single-cell Damagenome Profiling by Linear Copying and Splitting based Whole Genome Amplification (LCS-WGA)
    Authors:  Yichi Niu, Qiangyuan Zhu and Chenghang Zong, date: 03/20/2022, view: 522, Q&A: 0
    [Abstract]

    Spontaneous DNA damage frequently occurs on the human genome, and it could alter gene expression by inducing mutagenesis or epigenetic changes. Therefore, it is highly desired to profile DNA damage distribution on the human genome and identify the genes that are prone to DNA damage. Here, we present a novel single-cell whole-genome amplification

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    Phytophthora sojae Transformation Based on the CRISPR/Cas9 System
    Authors:  Jingting Cao, Min Qiu, Wenwu Ye and Yuanchao Wang, date: 03/20/2022, view: 570, Q&A: 0
    [Abstract]

    Phytophthora sojae is a model species for the study of plant pathogenic oomycetes. The initial research on gene function using Phytophthora was mainly based on gene silencing technology. Recently, the CRISPR/Cas9-mediated genome editing technology was successfully established in P. sojae and widely used in oomycetes. In this protocol, we describe

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    TUNEL Labeling to Detect Double-stranded DNA Breaks in Caenorhabditis elegans Gonads
    Authors:  Peter A. Kropp, Kyle Rhodehouse and Andy Golden, date: 03/20/2022, view: 433, Q&A: 0
    [Abstract]

    Analysis of DNA double strand breaks (DSBs) is important for understanding dyshomeostasis within the nucleus, impaired DNA repair mechanisms, and cell death. In the C. elegans germline, DSBs are important indicators of all three above-mentioned conditions. Although multiple methods exist to assess apoptosis in the germline of C. elegans, direct

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    Novel Protein Production Method Combining Native Expression in Human Cells with an Intein-based Affinity Purification and Self-cleavable Tag
    Authors:  Peter J. Carman and Roberto Dominguez, date: 03/20/2022, view: 488, Q&A: 0
    [Abstract]

    The human proteins used in most biochemical studies are commonly obtained using bacterial expression. Owing to its relative simplicity and low cost, this approach has been extremely successful, but is inadequate for many proteins that require the mammalian folding machinery and posttranslational modifications (PTMs) for function. Moreover, the

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