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  • So Do Vector

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    Trường Giang
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    The document describes the cloning of several genes into different plasmid vectors. The dUCH gene was cloned into pUAST and pBluescript vectors for expression in Drosophila and RNA interference studies. The dUCH and α-synuclein genes were also cloned into a pET28a vector for overexpression in E. coli using the T7 promoter. All cloning involved PCR amplification of the genes, digestion with restriction enzymes, and ligation into plasmids followed by transformation into E. coli DH5α and expression in BL21 cells.

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    The document describes the cloning of several genes into different plasmid vectors. The dUCH gene was cloned into pUAST and pBluescript vectors for expression in Drosophila and RNA interference studies. The dUCH and α-synuclein genes were also cloned into a pET28a vector for overexpression in E. coli using the T7 promoter. All cloning involved PCR amplification of the genes, digestion with restriction enzymes, and ligation into plasmids followed by transformation into E. coli DH5α and expression in BL21 cells.

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    © All Rights Reserved
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    9 views4 pages

    So Do Vector

    Uploaded by

    Trường Giang
    The document describes the cloning of several genes into different plasmid vectors. The dUCH gene was cloned into pUAST and pBluescript vectors for expression in Drosophila and RNA interference studies. The dUCH and α-synuclein genes were also cloned into a pET28a vector for overexpression in E. coli using the T7 promoter. All cloning involved PCR amplification of the genes, digestion with restriction enzymes, and ligation into plasmids followed by transformation into E. coli DH5α and expression in BL21 cells.

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    © All Rights Reserved
    Download as PPT, PDF, TXT or read online from Scribd
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    of 4

    pUAST/dUCH

    PCR product (dUCH gene)


    (1443bp)

    XhoI and BglII digestion

    T4 ligation

    -10339bp
    - have Amp resistant gene
    - For insertion of dUCH
    pUAST- dUCH
    gene to Drosophila
    chromosome
    - E. coli DH5α was used for
    cloning
    pBluescript/ NdUCH

    PCR product (NdUCH gene)


    (420bp)

    T4 DNA ligase

    -3381bp
    - have Amp resistant gene
    pBluuescript/NdUCH - For making dUCH dsRNA
    - E. coli DH5α was used for
    cloning
    pET/dUCH
    F1 origin

    PCR product (cDNA dUCH gene)


    (702bp)
    pET 28a
    kana lacI
    5369bp

    NdeI / EcoRI digestion

    F1 origin

    T4 DNA ligation
    - 6071bp
    kana pET 28a lacI - have Kanamycin
    NdeI/EcoRI
    dUCH resistant gene
    - For over expression
    kana dUCH gene in E. coli
    pET 28a - E. coli DH5α was used for
    dUCH
    cloning
    - E. coli BL21 (Dec3) was
    lacI
    used for over express
    pET/anpha- synuclein
    F1 origin

    Sản phẩm PCR gene anpha- synuclein


    (~400 bp)
    pET 28a
    kana lacI
    5369bp

    Xử lý bằng BamHI / BglI

    F1 origin

    Nối bằng T4 ligase


    - 5769 bp
    kana pET 28a lacI - have Kanamycin
    BamHI/BglI F1 origin
    Syn resistant gene
    - For over expression
    kana α-syn gene in E. coli
    pET 28a - E. coli DH5α was used for
    Synuclein
    cloning
    - E. coli BL21 (Dec3) was
    lacI
    used for over express

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