How does orcinol reagent for the estimation of RNA work?

HiPer® RNA Estimation Teaching Kit is designed for rapid and accurate determination of RNA by orcinol reagent. This method depends on the conversion of pentose in the presence of hot acid to furfural which then reacts with orcinol to yield a green colour. The colour intensity can be measured at 665 nm.

How is RNA quality determined?

The most common method used to assess the integrity of total RNA is to run an aliquot of the RNA sample on a denaturing agarose gel stained with ethidium bromide (EtBr). While native (non-denaturing) gels can be used, the results can be difficult to interpret.

Which test is used for quantification of RNA?

Quantification of messenger RNA (mRNA) by reverse transcription followed by nucleic acid amplification, usually polymerase chain reaction (PCR) [reverse transcriptase–quantitative PCR (RT–qPCR)], is widely performed, but the results are very dependent on the quality of the RNA.

Which colour produced when orcinol reacts with complex in RNA estimation?

green colour
Principle: The method usually involves the pentose conversion and ribose in the presence of hot acid to furfural that again reacts with orcinol to form a green colour.

What is orcinol reaction?

Rationale. When pentose is heated with concentrated hydrochloric acid, it forms furfural. Orcinol reacts with this furfural to give a green-colored compound. Ferric chloride acts as a catalyst.

What is the use of orcinol?

It is used in the production of the dye orcein and as a reagent in some chemical tests for pentoses, such as Bial’s Test.

How do you determine the quality of a RNA solution by using a spectrophotometer?

An A260 reading of 1,0 is equivalent to about 40 µg/ml of RNA and the OD at 260 nm is used to determine the RNA concentration in a solution. RNA has its absorption maximum at 260 nm and the ratio of the absorbance at 260 and 280 nm is used to assess the RNA purity of an RNA preparation.

What is RIN value?

The RNA integrity number (RIN) is an algorithm for assigning integrity values to RNA measurements. The integrity of RNA is a major concern for gene expression studies and traditionally has been evaluated using the 28S to 18S rRNA ratio, a method that has been shown to be inconsistent.

How do you test the purity of DNA and RNA?

To evaluate DNA purity, measure absorbance from 230nm to 320nm to detect other possible contaminants. The most common purity calculation is the ratio of the absorbance at 260nm divided by the reading at 280nm. Good-quality DNA will have an A260/A280 ratio of 1.7–2.0.

What is the basis of Bial’s orcinol test?

Principle of Bial’s Test This test is based on the principle that under hydrolysis pentosans are hydrolyzed into pentoses. Further, pentoses are dehydrated to yield furfural, which in turn condense with orcinol to form a blue-green precipitate.

What are the functions of orcinol and fecl3 in Bial’s test?

A method used to detect the presence of pentoses with a test reagent consisting of orcinol, HCl and ferric chloride. This test is used to detect the presence of a pentose in urine. In the presence of pentoses, the test reagent dehydrates pentoses to form furfural.

How do you make an RNA test with orcinol reagent?

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.

What is the reaction of RNA with orcinol?

But one of the widely used reactions is those of RNA with Orcinol. So, the basic objective of this test is to estimate RNA by orcinol method. Principle: The method usually involves the pentose conversion and ribose in the presence of hot acid to furfural that again reacts with orcinol to form a green colour.

What is orcinol method?

What is Orcinol Method? Orcinol method is a general reaction for pentoses. The orcinol reagent reacts with pentose groups in the backbone of the RNA molecule. Many downstream applications require high-quality RNA.

What is the reaction between orcinol and ferric chloride?

In the presence of ferric chloride as a catalyst, orcinol interacts with furfural to produce a green colour that can be detected at 665 nm. Standard RNA: Standard RNA solution- Dissolve 200µg/ml Standard RNA in 1 N perchloric acid/buffered Saline.