Ambion's Appendix - Base:Dye Ratio Calculator for the Silencer siRNA Labeling Kit

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Base:Dye Ratio Calculator

For the Silencer® siRNA Labeling Kit

Instructions
The approximate ratio of bases to dye molecules can be determined by measuring the absorbance of the labeled siRNA at 260 nm and at the maximum absorbance wavelength for the dye.

To perform these measurements, the labeled RNA should be at a concentration of at least 5 µg/ml (typically a 5-10 fold dilution of a Silencer™ siRNA labeling Kit reaction). See details.

Sample type:

ssRNA

dsRNA

Length of RNA (nt or bp):

Dye:

Notes About Calculation

Table 1. Reference Values for Spectrophotometry Calculations.

	Absorbance maximum	Extinction coefficient	Dye correction factor	MW_base
Cy3	550	150,000	0.08	1296
Cy5	649	250,000	0.05	1322
FAM	492	30,000	0.32	1006
21-mer ssRNA	260	9,700	––	320
21-mer dsRNA	260	8,000	––	320

A_base = (A₂₆₀) – (A_dye x dye correction factor)

base:dye ratio = (A_base x extinction coefficient_dye) / A_dye x extinction coefficient_base)

mg/ml nucleic acid = (A_base x MW_base) / (extinction coefficient_base x pathlength in cm)

where:

A_dye is the absorbance of the labeled nucleic acid at the dye's maximum absorbance wavelength
MW_base is the MW of a single nucleotide base
extinction coefficient_base is the extinction coefficient of a single nucleotide base

More information...

Ordering Information
For prices and availability, please contact our Customer Service Department.

Cat#	Product Name	Size
AM1632	Silencer® siRNA Labeling Kit - Cy™3	65 µg labeled siRNA
AM1634	Silencer® siRNA Labeling Kit - FAM™	65 µg labeled siRNA