Products

Fatty Tissue RNA Extraction Kit(SpinColumn)

Packing Specification：

Product Introduction：
The unique lysis buffer rapidly lyses cells and inactivates cellular RNases. After adjusting the binding conditions with ethanol, RNA is selectively adsorbed to the silica matrix membrane in the spin column under high chaotropic salt conditions. Through a series of rapid washing and centrifugation steps, deproteinization buffer and wash buffer remove impurities such as cellular metabolites and proteins. Finally, pure RNA is eluted from the silica matrix membrane with low-salt RNase-free H2O.

Product Features：
1.The silica matrix membrane in the spin column is made of high-quality special adsorption membrane, with minimal differences in adsorption capacity between columns.
2.No toxic reagents such as phenol or chloroform are used, and no ethanol precipitation steps are required.
3.Fast and simple: Operation for a single sample can generally be completed within 30 minutes.
4.Multiple column washing steps ensure high purity: The typical OD260/OD280 ratio ranges from 2.1 to 2.2 (The ratio of 100% pure RNA is generally around 2.2).

Application Scope：
Suitable for the rapid extraction of total RNA from tissue samples rich in fat, proteins, and extracellular substances.

Experimental Procedures：
Note:Before first use, add the indicated amount of absolute ethanol to the Wash Buffer RW bottle and 70% Ethanol bottle, mix thoroughly, and mark with a check in the box immediately after adding to avoid repeated addition!
1.For Fat-Rich Animal Tissue Samples
a.Liquid Nitrogen Grinding + Homogenization: Thoroughly grind the fat-rich tissue sample into fine powder in liquid nitrogen. Transfer an appropriate amount (40 mg) of the tissue fine powder into a 1.5 mL centrifuge tube containing 700 μL of Tissue Lysis Buffer RLT. Pipette to mix well more than 10 times with a 1 mL extended filter tip, or homogenize the lysate by drawing and expelling 10 times with a disposable 1 mL syringe (equipped with a 0.9 mm blunt needle) until a satisfactory homogenate is obtained (or homogenize electrically for 30 seconds). This can shear DNA, reduce viscosity, and improve yield.
b.Centrifuge the homogenized lysate at 13,000 rpm for 10 minutes at 2-8°C. The supernatant contains RNA. Remove insoluble substances in the homogenate: a large amount of fat floats on the top and must be removed; the remaining precipitate contains hard-to-lyse debris, extracellular membranes, and high-molecular-weight DNA. Carefully transfer the clear RNA-rich supernatant to a new centrifuge tube.
2.Accurately estimate the volume of the lysate (supernatant), aspirate the supernatant into a clean microcentrifuge tube, add 4 μL of Poly Carrier, pipette to mix well. Then add an equal volume of 70% ethanol (please check if absolute ethanol has been added first!). Precipitation may occur at this time, but it will not affect the extraction process. Immediately pipette to mix well; do not centrifuge.

3.Immediately transfer the mixed solution (less than 700 μL each time, can be added in two portions) to a SpinRA column (place the spin column into a collection tube), centrifuge at 13,000 rpm for 60 seconds, and discard the waste liquid.

4.Add 700 μL of Deproteinization Buffer RW1, incubate at room temperature for 30 seconds, centrifuge at 12,000 rpm for 30 seconds, and discard the waste liquid. If DNA residue is significant, incubate at room temperature for 5 minutes after adding RW1 before centrifugation.

5.Add 500 μL of Wash Buffer RW (please check if absolute ethanol has been added first!), centrifuge at 12,000 rpm for 30 seconds, and discard the waste liquid. Repeat this step with another 500 μL of Wash Buffer RW.

6.Place the SpinRA column back into the empty collection tube, centrifuge at 13,000 rpm for 2 minutes to remove as much wash buffer as possible, avoiding residual ethanol in the wash buffer that may inhibit downstream reactions.

7.Remove the SpinRA column, place it into a new RNase-free centrifuge tube. According to the expected RNA yield, add 30-50 μL of RNase-free water to the center of the adsorption membrane (heating the water in a 70-90°C water bath in advance can improve yield), incubate at room temperature for 1 minute, and centrifuge at 12,000 rpm for 1 minute.

8.If the expected RNA yield is >30 μg, add 30-50 μL of RNase-free water and repeat Step 8, then combine the two eluates. Alternatively, add the first eluate back to the spin column and repeat Step 8 for higher RNA concentration. 10 additional sets of RNase-free micro spin columns (SpinRA) and collection tubes are provided. For higher RNA concentration, elute RNA with 15-20 μL of RNase-free water.
Repeating the elution once results in a high-concentration RNA eluate. Combining two separate eluates increases the RNA yield by 15–30% compared to a single elution, but the concentration is lower. Users can choose according to their needs.