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    Protocols in Current Issue
    Single-step Precision Genome Editing in Yeast Using CRISPR-Cas9
    [Abstract] Genome modification in budding yeast has been extremely successful largely due to its highly efficient homology-directed DNA repair machinery. Several methods for modifying the yeast genome have previously been described, many of them involving at least two-steps: insertion of a selectable marker and substitution of that marker for the intended ...
    An Improved Method for Measuring Chromatin-binding Dynamics Using Time-dependent Formaldehyde Crosslinking
    [Abstract] Formaldehyde crosslinking is widely used in combination with chromatin immunoprecipitation (ChIP) to measure the locations along DNA and relative levels of transcription factor (TF)-DNA interactions in vivo. However, the measurements that are typically made do not provide unambiguous information about the dynamic properties of these ...
    Method for Multiplexing CRISPR/Cas9 in Saccharomyces cerevisiae Using Artificial Target DNA Sequences
    Authors:  Rachael M. Giersch and Gregory C. Finnigan, date: 09/20/2017, view: 4127, Q&A: 0
    [Abstract] Genome manipulation has become more accessible given the advent of the CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) editing technology. The Cas9 endonuclease binds a single stranded (single guide) RNA (sgRNA) fragment that recruits the complex to a corresponding genomic target sequence where it induces a double stranded ...
    Using CRISPR/Cas9 for Large Fragment Deletions in Saccharomyces cerevisiae
    Authors:  Huanhuan Hao, Jing Huang, Tongtong Liu, Hui Tang and Liping Zhang, date: 07/20/2017, view: 4016, Q&A: 0
    [Abstract] CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated protein 9) systems have emerged as a powerful tool for genome editing in many organisms. The wide use of CRISPR/Cas9 systems may be due to the fact that these systems contain a simple guide RNA (sgRNA) that is relatively easy to design and they are very ...
    Single-molecule Analysis of DNA Replication Dynamics in Budding Yeast and Human Cells by DNA Combing
    Authors:  Hélène Tourrière, Julie Saksouk, Armelle Lengronne and Philippe Pasero , date: 06/05/2017, view: 4649, Q&A: 0
    [Abstract] The DNA combing method allows the analysis of DNA replication at the level of individual DNA molecules stretched along silane-coated glass coverslips. Before DNA extraction, ongoing DNA synthesis is labeled with halogenated analogues of thymidine. Replication tracks are visualized by immunofluorescence using specific antibodies. Unlike biochemical ...
    CRISPR-PCS Protocol for Chromosome Splitting and Splitting Event Detection in Saccharomyces cerevisiae
    Authors:  Yu Sasano and Satoshi Harashima, date: 05/20/2017, view: 3719, Q&A: 0
    [Abstract] Chromosome engineering is an important technology with applications in basic biology and biotechnology. Chromosome splitting technology called PCS (PCR-mediated Chromosome Splitting) has already been developed as a fundamental chromosome engineering technology in the budding yeast. However, the splitting efficiency of PCS technology is not high ...
    Complex in vivo Ligation Using Homologous Recombination and High-efficiency Plasmid Rescue from Saccharomyces cerevisiae
    Authors:  Gregory C. Finnigan and Jeremy Thorner, date: 07/05/2015, view: 5343, Q&A: 0
    [Abstract] The protocols presented here allow for the facile generation of a wide variety of complex multipart DNA constructs (tagged gene products, gene fusions, chimeric proteins, and other variants) using homologous recombination and in vivo ligation in budding yeast (Saccharomyces cerevisiae). This method is straightforward, efficient ...
    Yeast DNA Replication 2D Gel Protocol
    Author:  Yanfei Zou, date: 07/05/2012, view: 9701, Q&A: 0
    [Abstract] Two-dimensional agarose gel electrophoresis (2D gel) analysis is used extensively as a method to detect origins of replication. Here, I present a simplified method for the isolation of yeast genomic DNA for 2D gel analysis from a small number of yeast cells. This DNA isolation method is simpler and less time consuming than the traditional method ...
    Gene Networks Based on the Graphical Gaussian Model
    Author:  Shisong Ma, date: 02/20/2012, view: 9199, Q&A: 1
    [Abstract] This protocol describes how to build a gene network based on the graphical Gaussian model (GGM) from large scale microarray data. GGM uses partial correlation coefficient (pcor) to infer co-expression relationship between genes. Compared to the traditional Pearson’ correlation coefficient, partial correlation is a better measurement of direct ...
    Illumina Sequencing Library Construction from ChIP DNA
    Author:  Wei Zheng, date: 02/20/2012, view: 14070, Q&A: 0
    [Abstract] The Illumina sequencing platform is very popular among next-generation sequencing platforms. However, the DNA sequencing library construction kit provided by Illumina is considerably expensive. The protocol described here can be used to construct high-quality sequencing libraries from chromatin immunoprecipitated DNA. It uses key reagents from ...



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