Flash content link: https://vlab.amrita.edu/repo/BIOTECH/MOL/HOT_SHOT_METHOD_DNA_EXTRACTION/index.swf

 

Requirements

 

•   Fins of Zebra fish
•   Micropipette and pipette tips
•   Scalpel
•   Micro centrifuge tubes
•   Mortar and pestle
•   Centrifuge
•   Water bath
•   4^oc freezer
•  -20^oc freezer

 

Reagents

 

•  50mM NaOH solution
•  1M Tris-Cl

 

Procedure

 

•  Pluck out 20-100 mg of fish fin using a sterile scalpel and a forceps.
•  Homogenize the fins using sterile mortar and pestle.
•  Aliquot 100 μl of 50 mM NaOH in to a microfuge tube.
• Transfer the homogenized fins of zebra fish into this micro centrifuge tube.
•  Incubate at 95°C for 20min in a water bath.
• After the incubation, the tubes are allowed to cool at 4°C.
•  Add 1/10th volume of 1 M Tris-HCl (pH 8.0) for neutralizing the basic solution.
• Centrifuge the sample at 5000rpm for 10min to pellet the debris. The supernatant obtained is collected in afresh micro centrifuge tube  (1-5μl of this supernatant can be used immediately  per 25 μL of  PCR reaction mixture).
• Store the sample for at least 3 months at 4°C or longer at -20°C freezer.
   

 

 

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