StarPure PAXgene Whole Blood Total RNA Extraction Kit

• The product can be stored stably for 18 months under appropriate storage conditions;
• This product can isolate and purify intercellular RNA from whole blood stored in PAXgene Blood RNA Tubes, forming an integrated system with sampling, stabilization and purification;
• Purification can be performed manually using a centrifuge or automatically on the QIAcube automated nucleic acid purification instrument.
• This product provides exceptional performance, ensuring highly reliable RNA purification.

Product components


Experimental procedures

1. PAXgene containing samples ^® Centrifuge the Blood RNA Tube at 3,000-5,000 × g for 10 min and discard the supernatant.

2. Add 4 mL of RNase-free water and vortex to thoroughly resuspend the pellet.

3. Centrifuge at 3,000-5,000 × g for 10 min and discard the supernatant.

✮Note: The supernatant should be removed as cleanly as possible, otherwise the lysis efficiency will be reduced, the lysate will be diluted, and the RNA yield will be reduced.

4. Add 350 µL of Resuspension Buffer to the pellet and vortex to thoroughly resuspend the pellet.

5. Transfer to a new centrifuge tube, add 300 µL Binding Buffer and 20 µL Proteinase K, and vortex for 5 seconds to mix.

6. Incubate on a 55°C heating shaker for 10 min.

7. Optional step: Aspirate the lysate at least five times with a syringe equipped with a 20-gauge needle (0.9 mm diameter) or process the lysate with an electric homogenizer to shear the genomic DNA, reduce the viscosity of the lysate, and increase the yield.

8. Centrifuge at 13,000 rpm for 3 min and transfer the supernatant to a new centrifuge tube.

9. Add 0.5× volume of anhydrous ethanol and vortex to mix.

10. Transfer 700 µL of the mixture to an RNA binding column (equipped with a 2 mL collection tube) and centrifuge at 13,000 rpm for 1 min.

11. Discard the liquid in the collection tube and place the adsorption column back into the collection tube.

12. Repeat steps 10-11 until all samples are transferred to the RNA adsorption column.

13. Add 350 µL Protein Wash Buffer and centrifuge at 13,000 rpm for 1 min.

14. Discard the liquid in the collection tube and place the adsorption column back into the collection tube.

15. Remove gDNA from the column and prepare the DNase I digestion reaction mixture as follows:

✮ Note :

• DNase I is very sensitive and easily denatured by physical means. Do not vortex the DNase I mixture. Invert the tube and mix gently.
• DNase I stock solution is prepared freshly for use.
• Standard DNase buffers are incompatible with on-membrane DNase I digestion. Using alternative buffers may affect RNA binding to the matrix and may reduce RNA yield and purity.
• All steps must be performed at room temperature, carefully and quickly.
• Pipette 50 µL of the DNase I digestion reaction mixture directly onto the membrane of the RNA binding column.

✮ Note : Be sure to pipette the DNase I digestion mixture directly onto the membrane. If some mixture remains on the walls or O-ring of the RNA binding column, the DNase I digestion will be incomplete.

17. Leave at room temperature for 15 minutes.

18. Add 350 µL Protein Wash Buffer and centrifuge at 13,000 rpm for 1 min.

19. Discard the liquid in the collection tube and place the adsorption column back into the collection tube.

20. Add 500 µL Wash Buffer and centrifuge at 13,000 rpm for 1 min.

21. Discard the liquid in the collection tube and place the adsorption column back into the collection tube.

22. Repeat steps 20-21.

23. Centrifuge again at 13,000 rpm for 2 min to remove residual liquid on the column.

✮ Note : Residual alcohol can affect downstream experiments.

24. Transfer the adsorption column to a new 1.5 mL centrifuge tube.

25. Add 50-70 μL RNase-free water (preheated to 70-90°C) to the membrane and let it stand at room temperature for 1 min.

26. Centrifuge at 13,000 rpm for 2 min, collect and properly store the eluted RNA sample.