One of the people in my lab added too much isopropanol to an RNA sample that was isolated with chloroform. What can I add to reduce the percent isopropanol? What is safe to add to an RNA sample/precipitation reaction? The sample was isolated with chloroform, so can I add chloroform to reduce the concentration of isopropanol?
Greetings,
You could just let it evaporate and resolubilize the pellet if you feel like, however an excess of IPA would just help you to maximize your RNA yield, as it dehydrates the nucleic acids present in your solution.
Best regards.
If you did your chloroform step right, you now have isopropanol mixed with the aqueous phase of the previous step. You should be ok if you add salt-containing buffer (ideally in the same concentration as your salt in the previous total volume) to make the volume of sample to isoprop 1:1. Important is that you need a minimum salt concentration, as the salt ions neutralize the phosphate groups of the RNA backbone, allowing efficient precipitation. You also need a minimum RNA concentration to ensure that a pellet can form, but I am assuming you have enough RNA.
Please refer for more information.
https://bitesizebio.com/2839/dna-precipitation-ethanol-vs-isopropanol/
Greetings İlknur Albayrak it's because the main purpouse for this is to fractionate the main components in solution, based on solubility. To the best of my knowledge and practice, as the protein precipitation methanol/chloroform, chloroform aims for proteins, while the main objetive of isopropanol is to dehydrate Nucleic acids.
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