TY - JOUR
T1 - Optimized protocol for the extraction of RNA and DNA from frozen whole blood sample stored in a single EDTA tube
AU - Yamagata, Hirotaka
AU - Kobayashi, Ayumi
AU - Tsunedomi, Ryouichi
AU - Seki, Tomoe
AU - Kobayashi, Masaaki
AU - Hagiwara, Kosuke
AU - Chen, Chong
AU - Uchida, Shusaku
AU - Okada, Go
AU - Fuchikami, Manabu
AU - Kamishikiryo, Toshiharu
AU - Iga, Jun ichi
AU - Numata, Shusuke
AU - Kinoshita, Makoto
AU - Kato, Takahiro A.
AU - Hashimoto, Ryota
AU - Nagano, Hiroaki
AU - Okamoto, Yasumasa
AU - Ueno, Shuichi
AU - Ohmori, Tetsuro
AU - Nakagawa, Shin
N1 - Funding Information:
This study was supported by the Japan Agency for Medical Research and Development (AMED) (JP20dk0307076h0003 to TO, SN, SU, and YO, and JP20dk0307075 to TAK). HY was partially supported by the Japan Society for the Promotion of Science (JSPS) KAKENHI program (20K07946); the Core Research for Evolutional Science and Technology (CREST); SENSHIN Medical Research Foundation; and grants from the Finding-Out & Crystallization of Subliminals (FOCS) project by Yamaguchi University of Medicine. The authors thank Dr. Kenji Hashimoto (Chiba University) for advice on blood sampling and RNA purification.
Funding Information:
HY has received research grants from Pfizer, Eisai, and MSD. SN has received honoraria and/or research grant support from Otsuka Pharmaceutical, Meiji Seika Pharma, Sumitomo Dainippon Pharma, Kyowa Pharmaceutical Industry, Shionogi, Mitsubishi Tanabe Pharma, Mochida Pharmaceutical, Eisai, Tsumura, Eli Lilly, MSD, Astellas, and Pfizer. TAK has received honoraria and/or research grant support from Otsuka Pharmaceutical, Mitsubishi Tanabe Pharma, Mochida Pharmaceutical, and Pfizer. TO has received research support or speakers’ honoraria from, or has served as a consultant to Sumitomo Dainippon Pharma, Otsuka Pharmaceutical, Eisai, Pfizer, Eli Lilly, Janssen, Meiji Seika Pharma, Shionogi, Mitsubishi Tanabe Pharma, Takeda Pharmaceutical, and Yoshitomiyakuhin. JI has received speaker’s honoraria from Otsuka Pharmaceutical, Meiji Seika Pharma, Sumitomo Dainippon Pharma, Kyowa Pharmaceutical Industry, Shionogi, Mochida Pharmaceutical, Eisai, Mylan, Sawai Pharmaceutical, Novartis Pharma, Eli Lilly, MSD, Ono Pharmaceutical, Takeda Pharmaceutical, Janssen Pharmaceutical, Sanofi, Viatris, and Yoshitomiyakuhin. The other authors declare no competing interests.
Publisher Copyright:
© 2021, The Author(s).
PY - 2021/12
Y1 - 2021/12
N2 - Cryopreservation of whole blood is useful for DNA collection, and clinical and basic research. Blood samples in ethylenediaminetetraacetic acid disodium salt (EDTA) tubes stored at − 80 °C are suitable for DNA extraction, but not for high-quality RNA extraction. Herein, a new methodology for high-quality RNA extraction from human blood samples is described. Quickly thawing frozen whole blood on aluminum blocks at room temperature could minimize RNA degradation, and improve RNA yield and quality compared with thawing the samples in a 37 °C water bath. Furthermore, the use of the NucleoSpin RNA kit increased RNA yield by fivefold compared with the PAXgene Blood RNA Kit. Thawing blood samples on aluminum blocks significantly increased the DNA yield by ~ 20% compared with thawing in a 37 °C water bath or on ice. Moreover, by thawing on aluminum blocks and using the NucleoSpin RNA and QIAamp DNA Blood kits, the extraction of RNA and DNA of sufficient quality and quantity was achieved from frozen EDTA whole blood samples that were stored for up to 8.5 years. Thus, extracting RNA from frozen whole blood in EDTA tubes after long-term storage is feasible. These findings may help advance gene expression analysis, as well as biomarker research for various diseases.
AB - Cryopreservation of whole blood is useful for DNA collection, and clinical and basic research. Blood samples in ethylenediaminetetraacetic acid disodium salt (EDTA) tubes stored at − 80 °C are suitable for DNA extraction, but not for high-quality RNA extraction. Herein, a new methodology for high-quality RNA extraction from human blood samples is described. Quickly thawing frozen whole blood on aluminum blocks at room temperature could minimize RNA degradation, and improve RNA yield and quality compared with thawing the samples in a 37 °C water bath. Furthermore, the use of the NucleoSpin RNA kit increased RNA yield by fivefold compared with the PAXgene Blood RNA Kit. Thawing blood samples on aluminum blocks significantly increased the DNA yield by ~ 20% compared with thawing in a 37 °C water bath or on ice. Moreover, by thawing on aluminum blocks and using the NucleoSpin RNA and QIAamp DNA Blood kits, the extraction of RNA and DNA of sufficient quality and quantity was achieved from frozen EDTA whole blood samples that were stored for up to 8.5 years. Thus, extracting RNA from frozen whole blood in EDTA tubes after long-term storage is feasible. These findings may help advance gene expression analysis, as well as biomarker research for various diseases.
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UR - http://www.scopus.com/inward/citedby.url?scp=85113775855&partnerID=8YFLogxK
U2 - 10.1038/s41598-021-96567-2
DO - 10.1038/s41598-021-96567-2
M3 - Article
C2 - 34426633
AN - SCOPUS:85113775855
SN - 2045-2322
VL - 11
JO - Scientific reports
JF - Scientific reports
IS - 1
M1 - 17075
ER -
