CRISPR-Cas9 sgRNA Library Design and Screening

  • DNA Editing
  • CRISPR-Cas9
    sgRNA Design
    sgRNA Library Construction
    sgRNA Panel
    Microbial Genome Editing
    Mammalian Cell Genome Editing
    ssDNA Synthesis
    CRISPR Amplicon Sequencing
    CRISPR NGS Data Analysis

    CRISPR-Cas9 technology has become a powerful tool for forward genetic screening. Mutant libraries of various functions can be constructed by CRISPR, and the genes related to this function then identified through functional screening and enrichment, PCR amplification, and deep sequencing analysis.

    Synbio Technologies provides one-stop gene function screening services based on the CRISPR-Cas9 sgRNA library system. With our proprietary Syno® 3.0 DNA synthesis platform, we can construct a sgRNA library in a fast and efficient manner. We can also package the library into lentivirus, transfect cells, perform high throughput/content screening, and finally identify the target genes by sequencing and data analysis.

    CRISPR-Cas9 sgRNA Library Process

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    sgRNA Library Construction and Screening Workflow

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    Competitive Advantages

    • Proprietary DNA synthesis platform: Fast and efficient sgRNA library construction
    • Lentivirus transfection system: Improve the efficiency of sgRNA transfection
    • Multiple screening platform: Including cell survival control, immunostaining, flow cytometry and other screening methods
    • One-stop solution: Customized services ranging from sgRNA library design, synthesis, and lentivirus packaging, to high throughput/content screening

    Service Specifications

    Service TypeService SpecificationsDeliverablesTAT
    CRISPR-Cas9 sgRNA lentivirus library (Human)Targeting 21,000 genes; 6 sgRNAs for each geneLentivirus, -80 ℃ preservationInquire
    Customized CRISPR-Cas9 sgRNA LibraryCustomized gRNA library, including genome-scale library, lncRNA library, cell apoptosis, cell proliferation, signal pathway, ion channel, nuclear receptor librariesLyophilized sgRNA plasmid sequencing report
    Lentivirus Packaging of sgRNA LibraryCustomized sgRNA design, 6 sgRNA for each gene Lentivirus packageLentivirus
    sgRNA Library ScreeningCell screening using cell survival control, immunostaining, flow cytometry and other screening methodsScreening report and sequencing data

    CRISPR-Cas9 sgRNA Library Design Case Study

    Screening resistance targets for a drug with identified function

    Methods

    1. The function and lethal dose of the drug have been already identified.
    2. CRISPR-Cas9 library was synthesized and transfected into cells using lentivirus.
    3. Screening: drug of high concentration acting on the mixed cell clone continuously over a short period, then screening the survived cells. These cells are considered to be resistant to this drug. The reason why these cells gain a resistance to this drug is due to mutagenesis caused by CRISPR-Cas9 gRNA.
    4. Perform NGS and bioinformatics analysis on the surviving cells, analyze information about the gRNA, and then identify the genes involved in conferring drug resistance.

    Results

    1. Identification of 6 genes involved in PLX resistant when mutant
    2. Identification of NF2 as the resistant target of PLX in melanoma

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    Reference

    Genome-Scale CRISPR-Cas9 Knockout Screening in Human Cells Ophir Shalem et al. Science 343, 84 (2014)