RNA and miRNA Yield from Whole Blood Samples Using Two Commercially
Available Isolation Methods
Church S.*, Schlueter K.**, Smith F.*
*BD Diagnostics PAS, Oxford, UK
**BD Diagnostics PAS, Heidelberg, Germany
Aim
RNA is very labile and must be stabilised immediately after blood collection to prevent
gene induction and RNA degradation. In this study we examined RNA and miRNA
yields from two commercial stabilisation and isolation methods.
Methods
In the first study RNA was extracted from 1378 samples using the PAXgene® Blood
RNA System, either manually or using the Qiagen QIAcube®. In a second study, blood
was collected from six subjects into PAXgene and Tempus RNA tubes. RNA was then
isolated using the Tempus™ Spin RNA Isolation Kit and the PAXgene Blood miRNA kit.
Three representative miRNA’s (mir-16; mir-30b; mir-103) were quantified in triplicate
using qRT-PCR performed on an ABI PRISM® TaqMan® 7900 System and
QuantiTect® SYBR® Green RT-PCR Kit. For both of these studies the isolation kits
use spin column isolation based methods.
Results
In the first study the RNA yield was ≥3 μg/2.5 mL blood in 99.8% of samples. In 82% of
samples the yield was 6–18μg and for all samples the mean yield was 9.3μg, (range
2.8-25 μg) which compares well with those published for the Tempus RNA tubes of 6-25
μg/3.0 mL blood. In the second study The CT for the three miRNA species stabilised
and purified with the two methods show that 3-20 times as much miRNA was isolated
from the PAXgene Blood miRNA System than the Tempus RNA kit.
Conclusion
Form these studies similar levels of performance were observed for RNA yield.
However there was a significant increase in the yield of miRNA using the PAXgene
Blood miRNA System when compared with the Tempus RNA kit.