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Protocol for Quick DNA Preparation for Genomic Fly PCR
Author: Jennifer Beck, updated date: , view: 58, Q&A: 0
Tags: DNA, Fly and Drosophila

Procedure

  1. Stock Solution:

    • 10 mM Tris-HCl (pH 8.2), 1 mM EDTA, 25 mM NaCl 

      • 100 ug/ml Proteinase K (add just before use) 

      • DNA extraction buffer: 

    • Proteinase K (10 mg/ml) 

  2. Method:

    • Put fly into 1.5 ml tube and keep on ice. 

    • Crush with 50 ul of DNA extraction buffer using pipet tip 

    • Centrifuge for 1 min at maximum speed 

    • Transfer supernatant in PCR tube 

    • Incubate for 30 min at 37°C 

    • Incubate for 5 min at 95°C 

    • Puton ice immediately 

    • Store at -20°C 

  3. PCR Protocol: 

    dH2O5.3 ul
    5x buffer2 ul
    dNTP (10mM)0.4 ul
    Primer A0.5 ul
    Primer B0.5 ul
    Tmplate1 ul
    Phusion Taq0.3 ul

    10 ul


  4. Cycler: 

    95°C2 min
    94°C0.5 min
    55°C2 min← x 30
    72°C2 min
    72°C5 min
    4°C


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