Improper preanalytical processes on peripheral blood compromise RNA quality and skew the transcriptional readouts of mRNA and LncRNA

He, Yinli1; Dong, Lele2; Yi, Hongyang3; Zhang, Linpei1; Shi, Xue1; Su, Lin1; Gan, Baoyu1; Guo, Ruirui1; Wang, Yawen1; Luo, Qinying1; Li, Xiaojiao1

2023-01-04

10.3389/fgene.2022.1091685

FRONTIERS IN GENETICS   Impact Factor & Quartile

1664-8021

Genetic and epigenetic reprogramming caused by disease states in other tissues is always systemically reflected in peripheral blood leukocytes (PBLs). Accurate transcriptional readouts of Messenger RNA (mRNA) and Long non-coding RNA (lncRNA) in peripheral blood leukocytes are fundamental for disease-related study, diagnosis and treatment. However, little is known about the impact of preanalytical variables on RNA quality and downstream messenger RNA and Long non-coding RNA readouts. In this study, we explored the impact of RNA extraction kits and timing of blood placement on peripheral blood leukocyte-derived RNA quality. A novel enhanced evaluation system including RNA yields, purity, RNA integrity number (RIN) values and beta-actin copies was employed to more sensitively identify RNA quality differences. The expression levels of informative mRNAs and Long non-coding RNAs in patients with chronic obstructive pulmonary disease (COPD) or triple-negative breast cancer (TNBC) were measured by Quantitative reverse transcription polymerase chain reaction (qRT-PCR) to investigate the impact of RNA quality on transcriptional readouts. Our results showed that the quality of RNA extracted by different kits varies greatly, and commercial kits should be evaluated and managed before batch RNA extraction. In addition, the quality of extracted RNA was highly correlated with the timing of blood placement, and the copy number of beta-actin was significantly decreased after leaving blood at RT over 12 h. More importantly, compromised RNA leads to skewed transcriptional readouts of informative mRNAs and Long non-coding RNAs in patients with chronic obstructive pulmonary disease or triple-negative breast cancer. These findings have significant implications for peripheral blood leukocyte-derived RNA quality management and suggest that quality control is necessary prior to the analysis of patient messenger RNA and Long non-coding RNA expression.

peripheral blood leukocytes (PBLs) preanalytical processes RNA quality transcriptional readouts COPD TNBC

SCI

English

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Science and Technology Planning Project of Shaanxi Province, China["2020KRM-202","2021JM-280","81903320"] ; National Natural Science Foundation of China[2021JM-280] ; institutional foundation of the first affiliated hospital of Xi'an Jiaotong University[2019QN-18] ; Fundamental Research Funds for the Central Universities[xzy012022112]

Genetics & Heredity

Genetics & Heredity

WOS:000922594800001

FRONTIERS MEDIA SA

Web of Science

1.Xi An Jiao Tong Univ, Affiliated Hosp 1, BioBank, Xian, Shaanxi, Peoples R China
2.Xi An Jiao Tong Univ, Affiliated Hosp 1, Dept Pharm, Xian, Shaanxi, Peoples R China
3.Southern Univ Sci & Technol, Peoples Hosp Shenzhen 3, Natl Clin Res Ctr Infect Dis, Affiliated Hosp 2, Shenzhen, Guangdong, Peoples R China

He, Yinli,Dong, Lele,Yi, Hongyang,et al. Improper preanalytical processes on peripheral blood compromise RNA quality and skew the transcriptional readouts of mRNA and LncRNA[J]. FRONTIERS IN GENETICS,2023,13.

He, Yinli.,Dong, Lele.,Yi, Hongyang.,Zhang, Linpei.,Shi, Xue.,...&Li, Xiaojiao.(2023).Improper preanalytical processes on peripheral blood compromise RNA quality and skew the transcriptional readouts of mRNA and LncRNA.FRONTIERS IN GENETICS,13.

He, Yinli,et al."Improper preanalytical processes on peripheral blood compromise RNA quality and skew the transcriptional readouts of mRNA and LncRNA".FRONTIERS IN GENETICS 13(2023).