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  • Development of A Plant Promoter: Gh-Rbcs Promoter - Leaves Specific

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    Chandra Prakash
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    The document describes the process of developing a plant promoter. Genomic DNA was isolated from leaves and digested to select a fragment containing the Gh-rbcS gene sequence. Primers were designed to amplify the gene via PCR. Linkers were added to the PCR product, which was then digested and ligated into a binary vector. The vector was transformed into Agrobacterium and used to transform plants. GUS expression was observed in leaves, confirming the leaf-specific Gh-rbcS promoter.

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    Promoter

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    The document describes the process of developing a plant promoter. Genomic DNA was isolated from leaves and digested to select a fragment containing the Gh-rbcS gene sequence. Primers were designed to amplify the gene via PCR. Linkers were added to the PCR product, which was then digested and ligated into a binary vector. The vector was transformed into Agrobacterium and used to transform plants. GUS expression was observed in leaves, confirming the leaf-specific Gh-rbcS promoter.

    Copyright:

    Attribution Non-Commercial (BY-NC)
    Download as PPT, PDF, TXT or read online from Scribd
    Flag for inappropriate content
    Download as ppt, pdf, or txt
    10 views5 pages

    Development of A Plant Promoter: Gh-Rbcs Promoter - Leaves Specific

    Uploaded by

    Chandra Prakash
    The document describes the process of developing a plant promoter. Genomic DNA was isolated from leaves and digested to select a fragment containing the Gh-rbcS gene sequence. Primers were designed to amplify the gene via PCR. Linkers were added to the PCR product, which was then digested and ligated into a binary vector. The vector was transformed into Agrobacterium and used to transform plants. GUS expression was observed in leaves, confirming the leaf-specific Gh-rbcS promoter.

    Copyright:

    Attribution Non-Commercial (BY-NC)
    Download as PPT, PDF, TXT or read online from Scribd
    Flag for inappropriate content
    Download as ppt, pdf, or txt
    of 5

    Development of a plant promoter

    Gh-rbcS promoter Leaves specific

    Genomic DNA isolation, Purification Hind III digestion- Agarose gel electrophoresis Selected fragment elution Genbank- Specific gene sequence Regulatory sequences oligonucleotide design (Primers) for PCR gene amplification

    Ex: Gh-rbcS gene


    5'

    3'

    5' primer
    5'-cgctcatgtt aacaattaat tcctataatc-3'

    3 primer
    5'-catcgtagta cgtgggtaag ctcgagtact-3'

    PCR products
    5' 3'

    Addition of Linker to the 5 end


    5' PCR mixture 5' 3' 3'

    PCR products 5' 3' 5'

    Nco I

    3'

    Pst I

    Digestion with Pst I and Nco I Ligated into the Pst I and Nco I sites in the binary vector pCGN1578

    Gh rbcS/GUS/CaMV 35S term-CaMV35 pro/NPT/CaMV 35S ter

    Binary vector inserted into Agrobacterium tumefaciens strain EHA101 Transform the plant with Agrobacterium. GUS assay- GUS expression in leaves

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