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    Home > Biochemistry News > Microbiology News > JGG Huazhong Agricultural University's Peng Nan team and He Mingxiong team developed an efficient editing tool for green bio-manufacturing chassis cells

    JGG Huazhong Agricultural University's Peng Nan team and He Mingxiong team developed an efficient editing tool for green bio-manufacturing chassis cells

    • Last Update: 2021-11-15
    • Source: Internet
    • Author: User
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    Recently, the State Key Laboratory of Agricultural Microbiology of Huazhong Agricultural University and the research group of Professor Nan Peng from Hongshan Laboratory of Hubei Province and the team of He Mingxiong Research Institute of Biogas Science Research Institute of the Ministry of Agriculture and Rural Affairs revealed the working rules of the new bacterial DNA damage repair system.
    Combining with the bacterial endogenous CRISPR-Cas system, an efficient genome editing tool for the important chassis of green organisms, Zymomonas mobilis and Pediococcus lactis has been established
    .

    Figure 1 Zymomonas mobilis uses the MMEJ pathway to differentially repair the damage of genomic DNA and movable elements caused by the CRISPR-Cas system.
    Ethanol-producing bacteria have the characteristics of high ethanol fermentation rate, good ethanol tolerance and biosafety, and have great application value in biofuel applications
    .

    The research team found that the genomic DNA damage caused by the CRISPR-Cas system can be efficiently repaired by the micro-homology-mediated joining pathway (MMEJ) and cause partial DNA deletion
    .

    The MMEJ system has a very low efficiency in repairing the damage of moving elements caused by CRISPR-Cas, which represents a recognition mode that distinguishes itself from foreign DNA after cutting
    .

    The result was published as a cover article in the Journal of Genetics and Genomics (Figure 1).
    Doctoral student Xiaojie Wang from Huazhong Agricultural University is the co-first author of the paper
    .

    Figure 2 The rapid genome editing process based on the endogenous CRISPR-Cas system and the MMEJ pathway.
    On this basis, the research team used the endogenous CRISPR-Cas system and the MMEJ pathway to develop a rapid genome suitable for Zymomonas mobilis Tools for editing and identification of essential genes
    .

    The tool only needs to construct a self-targeting plasmid with mini-CRISPR (Repeat-Spacer-Repeat) to achieve the knockout of target genes and long fragments (Figure 2)
    .

    In addition, the editing efficiency of the tool is further improved by modifying the second repeat sequence (Repeat) of mini-CRISPR
    .

    The results were published in the journal Biotechnology for Biofuels, and Dr.
    Sui Xin from Huazhong Agricultural University was the first author of the paper
    .

    Figure 3 The integrated expression of lactate dehydrogenase gene significantly improves the growth rate and lactic acid production rate of Pediococcus lactis.
    Pediococcus lactis is an important probiotic in the feed and medicine catalogs.
    It is widely used in lactic acid and feed fermentation and immune regulation and other production practices and In clinical treatment
    .

    The research team isolated a strain of Pediococcus lactis with good production performance and strong probiotic properties in the early stage, and found that its genome encodes the CRISPR-Cas9 system
    .

    Although the cas9 gene has a nonsense mutation, research by the research team has shown that it can still perform the function of nucleic acid cleavage
    .

    Based on the endogenous CRISPR-Cas9 system, the research team has established efficient genetic manipulation tools to achieve efficient gene knockout, point mutation and integration
    .

    Using this technology to eliminate endogenous plasmids and integrate over-expressed lactate dehydrogenase genes significantly increased the growth rate of Pediococcus lactis and the rate of lactic acid production (Figure 3)
    .

    The results were published in the Journal of Applied and Environmental Microbiology.
    Liu Ling, a PhD student from Huazhong Agricultural University, was the first author of the paper, and the research team was granted an invention patent
    .

    This work has laid a solid foundation for the research on synthetic biology of probiotics
    .

    Article link: https://doi.
    org/10.
    1016/j.
    jgg.
    2021.
    02.
    012https://doi.
    org/10.
    1186/s13068-021-02056-zhttps://doi.
    org/10.
    1128/AEM.
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