A comprehensive, cell specific microRNA catalogue of human peripheral blood

Abstract

With this study, we provide a comprehensive reference dataset of detailed miRNA expression profiles from seven types of human peripheral blood cells (NK cells, B lymphocytes, cytotoxic T lymphocytes, T helper cells, monocytes, neutrophils and erythrocytes), serum, exosomes and whole blood. The peripheral blood cells from buffy coats were typed and sorted using FACS/MACS. The overall dataset was generated from 450 small RNA libraries using high-throughput sequencing. By employing a comprehensive bioinformatics and statistical analysis, we show that 3′ trimming modifications as well as composition of 3′ added non-templated nucleotides are distributed in a lineage-specific manner—the closer the hematopoietic progenitors are, the higher their similarities in sequence variation of the 3′ end. Furthermore, we define the blood cell-specific miRNA and isomiR expression patterns and identify novel cell type specific miRNA candidates. The study provides the most comprehensive contribution to date towards a complete miRNA catalogue of human peripheral blood, which can be used as a reference for future studies. The dataset has been deposited in GEO and also can be explored interactively following this link: http://134.245.63.235/ikmb-tools/bloodmiRs.

qPCR-Based Methods for Expression Analysis of miRNAs

“…The PanelChip™ Analysis System was developed and can contain up to 2500 assays, being based on the use of nanowells and qPCR [66]. In recent years, approaches based on RNA sequencing (RNA-seq), which use methods for massive sequencing (such as the ones from Illumina or SOLiD), have shown their large potential for identifying the complete profiles of miRNA variability in several cells and tissues [40,44,60,67]. …”

“…There are several freely available programs that are helpful for several steps in the analysis of miRNA expression, including miRbase [5], TarBase [48], a cell-specific microRNA catalogue of human peripheral blood [40], miRTarBase [49], miRmine [50], UCSC Genome Browser [51], Ensembl Genome Browser [52], Primer3 [53], BatchPrimer3 [54], miRPrimer [55], RTPrimerDB [56] and mfold [57] (Table 1). …”

Depletion of erythropoietic miR-486-5p and miR-451a improves detectability of rare microRNAs in peripheral blood-derived small RNA sequencing libraries

“…Compositional analysis of miRNA species in blood compounds was performed using processed miRNA count data from our previously published study (10). The dataset contains miRNA expression counts of seven types of blood cells, exosomes, serum and whole blood, which were generated by using TruSeq (Illumina) small RNA library preparation protocol.…”

“…The oligos are designed to prevent 5' adapter ligation via blocking the access of T4 RNA ligase to the 5' end of target miRNAs during smRNAseq library preparation. Sequences of two blocking oligos were composed based on our previously published whole blood smRNA-seq data (10). Briefly, all sequences that were mapped to precursors of miR-486-5p and miR-451a were pooled in order to obtain all possible unique sequence variants for each miRNA.…”

“…To test this hypothesis, we performed a compositional analysis of miRNA species in blood compounds and whole blood samples using blood miRNA catalog data (10). Here, for each sample, we calculated the Shannon diversity index, which combines the information about miRNA richness (number of detected different miRNA species) and evenness (proportion of each miRNA) within a given sample (25).…”