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        Midiprep preparation of Plasmid DNA

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        1662
        实验概要


        The PureLink™ HiPure Plasmid DNA Midiprep Kit allows purification of 100–350 μg of high-quality plasmid DNA from 15–25 mL overnight E. coli cultures in ~2 hours when cloning high copy number plasmids.

        主要 试剂

        Before Starting
        Before beginning, verify that the Resuspension Buffer (R3) contains RNase A, and no precipitate has formed in the Lysis Buffer (L7).
        Materials Needed
        Overnight culture of transformed E. coli cells
        Isopropanol
        70% ethanol
        Sterile, microcentrifuge tubes
        PureLink™ Nucleic Acid Purification Rack
        Tubes or centrifuge bottles for harvesting cells
        Centrifuge and rotor appropriate for harvesting cells
        Appropriate 15-mL centrifuge tubes capable of
        withstanding centrifugation forces >12,000 × g
        Centrifuge capable of centrifuging at >12,000 × g at 47°C
        Optional : PureLink™ HiPure Precipitator Module
        Components Supplied with the Kit
        Resuspension Buffer (R3) with RNase A
        Lysis Buffer (L7)
        Precipitation Buffer (N3)
        Equilibration Buffer (EQ1)
        Wash Buffer (W8)
        Elution Buffer (E4)
        TE Buffer (TE)
        HiPure Filter Midi Columns
        Column Holder
        Note: The protocol for the Midiprep kit of the PureLink™ HiPure Plasmid Filter Purification Kits is designed for purification of both high and low copy number plasmids without the need for adjusting buffer volumes.

         

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