Bio-Rad Aurum™ Total RNA Fatty and Fibrous Tissue Kit - Manual (Page 9)

5
Section 5
Before Using the Aurum™ Total RNA
Fatty and Fibrous Tissue Kit
Please read the following guidelines before proceeding with the total RNA
isolation.
Maximum Starting Material Amounts*
The Aurum total RNA fatty and fibrous tissue kit is designed to process up to
the amounts indicated below (per column):
• 1 x 10 7 cultured mammalian cells grown in suspension
• One 10 2 cm plate mammalian cultured cells grown in monolayer
• 2.4 x 10 9 of gram-positive or gram-negative bacteria (equivalent to
3 OD•ml of bacteria)
• 3.0 x 10 7 of yeast (equivalent to 3 OD•ml of yeast)
• 100 mg of animal tissue (a 4 mm cube of most animal tissue weighs
approximately 70–85 mg)
• 100 mg of plant tissue
• 50 mg filamentous fungi
Warning: Processing larger amounts of starting material may lead to column
clogging and reduced RNA purity. It is crucial that the appropriate amount of
starting material be used. For samples that are known to be rich in RNA, it is
highly recommended that less than the maximum amount of starting material
be used so that the binding capacity of the column is not exceeded. In
addition, complete disruption and homogenization of the starting material is
critical to prevent column clogging and reduced RNA yields.
Minimum Starting Material Amounts
• 50 cultured mammalian cells
• 5 mg of animal tissue
• 5 mg plant tissue
* Spectrophotometric determination of bacterial or yeast culture density is a
REQUIREMENT for optimal total RNA isolation from these starting materials.
To determine the density of a bacterial or yeast culture (OD
600
), combine
50 µl of culture with 950 µl growth medium (20-fold dilution). Use the growth
medium as a blank and take the spectrophotometric reading at 600 nm.
Multiply this figure by 20 to calculate the OD
600
value of the undiluted
bacterial or yeast culture. Depending upon the OD
600
value, a specific volume
of the culture will be selected to provide an optimum amount of bacteria or
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