Substitution of Chloroform with 1-Bromo-3-chloropropane in RNA-isolation.
Chloroform is commonly used in a method to isolate RNA from DNA and proteins. Since chloroform is a neurotoxicant, an endocrine disruptor and possibly also a carcinogen, substitution of the substance is advantageous from a workers safety perspective. It was found that 1-Bromo-3-chloropropane (BCP) could work as a fully functional substitute for chloroform without any changes in the experimental process.
Chloroform is in the database of hazardous substances according to SUBSPORT screening criteria (SDSC), while the alternative is not. Chloroform holds the risk phrases R22, 38, 40 and 48/30/22. The alternative holds R22 and R40.
In phenol-chloroform extraction, RNA is separated from DNA and proteins via a 2-step procedure. (The same procedure can also be used to separate DNA or proteins from the other components.)
Chloroform is used in the first step where a mixture of chloroform and phenol is mixed with an aqueous solution together with the sample containing DNA, RNA and proteins. The combined mixture is then centrifuged which results in one organic phase and one inorganic phase. The inorganic phase, containing both nucleic acids will be placed on top of the vessel allowing it to be extracted. The organic phase, mainly consisting of chloroform will be placed at the bottom. This phase will contain the proteins of the mixture.
DNA can then be separated from RNA in a further step using 2-propanol or ethanol. The substitution of chloroform with BCP does not include any changes in the experimental process.
BCP is a less hazardous solvent than chloroform which results in safer working place conditions mainly for the laboratory assistant.
The alternative is a less hazardous substance than chloroform, which is of advantage mainly for the laboratory personnel. However due to the properties of the alternative “harmful if swallowed” and “limited evidence of carcinogenic effect” the alternative should be handled carefully.
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Laboratory assistant at Sahlgrenska University Hospital, department of Clinical Chemistry.
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