Bio-Rad SmartSpec Plus Spectrophotometer - Manual (Page 30)

b. If you do not know the molecular weight.
• Use the number keys to input the number of A’s, then press Enter to move the
cursor to the next field and enter the number of C’s. After entering the number
of T’s (or U’s for RNA oligos), press Enter. SmartSpec Plus will calculate the
molecular weight and display it along with the molar extinction coefficient.
• Press the right arrow key to continue the assay. SmartSpec Plus is now ready
to collect absorbance measurements and to convert those to concentrations.
ii. If you do not know the molar extinction coefficient, but you know the
mass/absorbance conversion factor. If you do not know the molar extinction
coefficient, SmartSpec Plus will ask for the mass/absorbance conversion factor.
This factor would be multiplied by the absorbance to calculate mass concentration
(e.g., for dsDNA the mass/absorbance conversion factor is 50.0 µg/ml for an
A260 of 1.0). If you do not know either the molar extinction coefficient or the
mass/absorbance conversion factor, SmartSpec Plus will estimate them for you.
• If you know the conversion factor, then press Select to toggle from No to Yes and
press Enter. If you do not know the conversion factor, press Enter and skip down
to iii.
• Use the number keys to input the conversion factor and press Enter. SmartSpec
Plus will now need the molecular weight of the oligonucleotide in order to convert
absorbance measurements into concentrations of µg/ml and pmol/µl.
• If you know the molecular weight, press Select to toggle from No to Yes and press
Enter. If you do not know the molecular weight of the oligo, press Enter and skip
to b below.
a. If you know the molecular weight.
• Use the number keys to input the molecular weight and press Enter.
After you have entered the molar extinction coefficient and the molecular weight,
SmartSpec Plus will display them.
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Mol Ext Coef=xxxxxxx
MW=xxxxx >=Continue
Enter oligo composition:
#A:___#C:___#G:___#T:___
Do you know conv factor
from A260 to conc? No
Mole Ext Coef=xxxxxxx
MW=xxxxx >=Continue
Enter oligo molecular
weight: _________ g/mol
Enter conversion factor:
A260 1.0 = ________ µg/ml
Do you know molecular
weight of oligo? No