Estimation of RNA by the Orcinol Method

Principle
This is a general reaction for pentose and depends on the formation of furfural. When pentose is heated with concentration HCl, orcinol reacts in presence of furfural in presence of ferric chloride as a catalyst purine to produce green color only the purine nucleotide.

Materials

• RNA commercial sample: 10 mg
• RNA solution (0.2 mg/mL): Dissolve 10 mg of commercially available RNA in 50 mL of buffer saline and use the sample for standard preparation.
• Orcinol reagent: Dissolve 10 gm of ferric chloride (FeCl₃)6 H₂O in 1 liter of 1 gm of ferric chloride (FeCl)6 H₂O in 1 liter of concentrated HCl and add 35 mL of 6% w/v orcinol in alcohol.
• Buffered saline.
• Boiling water bath.

Procedure
Pipette out standard RNA solution in arrange of 0–2 mL into a series of test tube and make up the volume of each tube 2 mL with distilled water. Add 3 mL of orcinol reagent to each tube for standard solution. For test solution : Take 2 mL of nucleic acid sodium (isolated from tissue source.) Add 3 mL of orcinol reagent to each tube, and heat the tube on boiling water bath for 20 minutes. Cool and take the optical density at 665 nm against the orcinol blank.

Result:
Concentration of RNA present in the given sample is .............. µg/2 mL.