Team:HKUST/Protocols/Ethanol precipitation

Ethanol precipitation

Purpose: To concentrate DNA product.

Materials: 100% ethanol, 75% ethanol, 3M NaAc, ddH2O, DNA product

Procedure:

a.Add 2 volume of 100% ethanol.
b.Add 0.1 volume of 3M NaAc.
c.Put the tube in fridge at -20 °C for 10 minutes.
d.Put it out and centrifuge it for 10-15 minutes (13,000 rpm). Remove the supernatant.
e.Wash with 100μL 75% ethanol.
f.Centrifuge for 5 minutes (13,000 rpm). Remove all the supernatant.
g.Add 10μL ddH2O to resuspend DNA.

Tips:

A.If the volume of DNA product is too low, make it to higher volume with ddH2O. The recommended lowest volume is 50μL. Combine several tubes of DNA product is also suggested for higher DNA product concentration.
B. In step f, make sure to remove all the supernatant without touching the pellet. If it is too hard to do so, open the tube and leave it at room temperature for a while to make the ethanol evaporate.

• Agarose gel preparation and gel electrophoresis
• PCR (Taq, Vent)
• PCR product clean-up
• Enzyme digestion (vector, insert)
• Plasmid DNA extraction
• Gel cutting
• Gel extraction
• Ethanol precipitation
• Ligation
• Transformation to E.coli
• Transformation to yeast
• Cell lysis
• Yeast genomic DNA extraction
• Western blotting
• GFP testing