Hello, not so big problem greed with above answers only thing for better isolation put in freezer for 15 min before centrifugation.

Good luck!

yes you can proceed with the process for RNA isolation, Use ethanol in all the steps instead of Isopropanol. No need to worry about.

• Agreed
• Add 2-3 volumes of ethanol for DNA extraction and > 3 volumes of ethanol for RNA extraction
• The only difference is that when you add 0.8-1.0 volumes of room temp isopropanol you spin immediately whereas for RNA isolation you need to incubate in 3 volumes of ethanol @ -20C for 1 hour before spinning
• Then everything else is the same as isopropanol procedure
• So 3 volumes instead of 3 and incubate in freezer before spinning; unlike iso

Hello, not so big problem greed with above answers only thing for better isolation put in freezer for 15 min before centrifugation.

Good luck!

• Not sure I completely agree with Karens answer
• Text book generally state 15 min @ -20C up to 1 hour @ -20C
• In general the longer you leave @ -20C the better the yield but the more likely you are to precipitate salt as well as RNA
• This is why you do not cool at all with isopropanol as salt more readily precipitates from chilled isopropanol compared with ethanol
• Thus 15 min is meant to represent the ambient balance between yield and co ppt of salt
• Whilst it is true that 5 min of chilling will be much better than no chilling at all - and you will precipitate RNA and for that duration probably free of salt - you might not precipitate all of your RNA; This is obviously an issue if you have low amounts of starting material and therefore anticipated low yields of RNA: Typically < 5ug total (in say 50ul)
• This if you are extracting from whole flasks of TC cells or grams of tissue 5min will be fine and will give you enough RNA which should be free of salt
• For smaller amounts of starting material however I would recommend a minimum of 15 min @ -20C. This incidentally is especially true of RNA compared to DNA