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W F Yang, Y H Tian, T T Wang

    Tsitologiia I Genetika
    |November 29, 2018
    PubMed
    Summary
    This summary is machine-generated.

    We developed a MuDR-TAIL-PCR system to efficiently isolate MuDR insertion-specific flanking sequences (MuDRFs). This method aids in utilizing Mu element-mediated mutants for genetic research.

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    Area of Science:

    • Molecular Biology
    • Genetics
    • Plant Science

    Background:

    • MuDR elements show high transposition activity in the Mutator (Mu) family.
    • Isolating MuDR insertion-specific flanking sequences (MuDRFs) is crucial for utilizing Mu element-mediated mutants.

    Purpose of the Study:

    • To construct and optimize a MuDR-TAIL-PCR system for specifically isolating MuDRFs.
    • To assess the efficiency and accuracy of the developed system in identifying MuDR insertions.

    Main Methods:

    • Developed a MuDR-TAIL-PCR system using MuDR-TIR-nested specific primers and arbitrary degenerate primers.
    • Optimized reaction conditions and procedures using mutant DNA templates from 87 genotypes.
    • Applied the system to isolate MuDRFs from M2 or M2:3 families.

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    Main Results:

    • Successfully acquired 129 different MuDRFs using the MuDR-TAIL-PCR system, representing 86.60% of total mutant-specific bands.
    • Confirmed the authenticity of non-redundant flanking sequence amplifications, accounting for 65.12% of total MuDRFs.
    • Found that 88.00% of non-redundant MuDRFs were inserted within genes, indicating high target specificity.

    Conclusions:

    • The developed MuDR-TAIL-PCR system is effective for specifically isolating MuDR flanking sequences.
    • This method facilitates the characterization and utilization of Mu element-induced mutations.
    • The system demonstrates high efficiency in identifying gene-inserted MuDR elements.