DNA lab report

DNA lab report - Materials and Methods Isolation of...

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Materials and Methods Isolation of chromosomal DNA from Vibrio fischeri. Centrifuge culture to isolate bacteria. Treat isolated bacteria with TES buffer to destabilize outer membrane. Add Lysozyme to break down peptidoglycan and let cool on ice. Add Proteinase K and incubate at 55c for 10 min to degrade periplasmic nucleases. Add SDS and incubate at 55c for 15 min to disrupt membranes and to denature proteins. Add Tris-buffered phenol to denature proteins then centrifuge to isolate DNA from precipitated proteins. Add 95% ethanol to precipitate DNA then let dry before inserting it in tube of TE. Add RNase and incubate for at 45c for 30 min then add Proteinase K to inactivate RNase. Add Phenol:Chloroform and Centrifuge for 5min. Collect top aqueous layer and repeat centrifuge using just 1 part Chloroform. Add sodium acetate and 95% ethanol and store on ice for 10min. Collect DNA with glass hook and let dry Make 1:100 dilution of DNA sample and run spectrophotometer to verify isolation of
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DNA lab report - Materials and Methods Isolation of...

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