Isolating high-quality RNA for RNA-Seq from 10-year-old blood samples

Abstract

There is much interest in analysing RNA, particularly with RNA Sequencing, across both research and diagnostic domains. However, its inherent instability renders it susceptible to degradation. Given the imperative for RNA integrity in such applications, proper storage and biobanking of blood samples and successful subsequent RNA isolation is essential to guarantee optimal integrity for downstream analyses. Especially for larger collections, it would be particularly beneficial if these methods would additionally offer affordability, minimal blood volume requirements and also long-term storage. In this study, RNA of high quality, suitable for transcriptomics, has been successfully isolated from 400 μL of EDTA and citrated whole blood samples in Boom’s lysis buffer stored at −85 °C for 10 years. Isolation was carried out using a modified Zymo Research Quick-RNA kit protocol. This isolation method showed significant improvement in RNA integrity when compared to RNA extracted using the original Boom method. RNA Sequencing provided high-quality data comparable to that of other studies using recently frozen blood in RNA stabilisation tubes. Additionally, sequencing data from blood collected in citrate and EDTA anticoagulants also showed excellent correlation.