tetano
Editor, Senior Moderator
Sci Rep
. 2024 Nov 5;14(1):26791.
doi: 10.1038/s41598-024-75740-3. Data normalization of plasma miRNA profiling from patients with COVID-19
Julia Tiemi Siguemoto[SUP] #[/SUP][SUP] 1 [/SUP], Carolini Motta Neri[SUP] #[/SUP][SUP] 1 [/SUP], Nadine de Godoy Torso[SUP] 2 [/SUP], Aline de Souza Nicoletti[SUP] 2 [/SUP], Marília Berlofa Visacri[SUP] 3 [/SUP], Carla Regina da Silva Correa da Ronda[SUP] 1 [/SUP], Mauricio Wesley Perroud Jr[SUP] 2 [/SUP], Leonardo Oliveira Reis[SUP] 2 [/SUP], Luiz Augusto Dos Santos[SUP] 4 [/SUP], Nelson Durán[SUP] 2 [/SUP], Wagner José Fávaro[SUP] 2 [/SUP], Eder de Carvalho Pincinato[SUP] 2 [/SUP], Patricia Moriel[SUP] 5 [/SUP]
Affiliations
When using the reverse-transcription quantitative polymerase chain reaction (RT-qPCR) technique for quantitative assessment of microRNA (miRNA) expression, normalizing data using a stable endogenous gene is essential; however, no universally adequate reference gene exists. Therefore, in this study, we aimed to determine, via the RNA-Seq technique, the most adequate endogenous normalizer for the expression assessment of plasma miRNAs in patients with coronavirus disease 2019 (COVID-19). Two massive sequencing procedures were performed (a) to identify differentially expressed miRNAs between patients with COVID-19 and healthy volunteers (n = 12), and (b) to identify differentially expressed miRNAs between patients with severe COVID-19 and those with mild COVID-19 (n = 8). The endogenous normalizer candidates were selected according to the following criteria: (1) the miRNA must have a fold regulation = 1; (2) the miRNA must have a p-value > 0.990; and (3) the miRNAs that were discovered the longest ago should be selected. Four miRNAs (hsa-miR-34a-3p, hsa-miR-194-3p, hsa-miR-17-3p, and hsa-miR-205-3p) met all criteria and were selected for validation by RT-qPCR in a cohort of 125 patients. Of these, only hsa-miR-205-3p was eligible endogenous normalizers in the context of COVID-19 because their expression was stable between the compared groups.
Keywords: COVID-19; Endogenous normalizer; MicroRNA; Reference gene.
. 2024 Nov 5;14(1):26791.
doi: 10.1038/s41598-024-75740-3. Data normalization of plasma miRNA profiling from patients with COVID-19
Julia Tiemi Siguemoto[SUP] #[/SUP][SUP] 1 [/SUP], Carolini Motta Neri[SUP] #[/SUP][SUP] 1 [/SUP], Nadine de Godoy Torso[SUP] 2 [/SUP], Aline de Souza Nicoletti[SUP] 2 [/SUP], Marília Berlofa Visacri[SUP] 3 [/SUP], Carla Regina da Silva Correa da Ronda[SUP] 1 [/SUP], Mauricio Wesley Perroud Jr[SUP] 2 [/SUP], Leonardo Oliveira Reis[SUP] 2 [/SUP], Luiz Augusto Dos Santos[SUP] 4 [/SUP], Nelson Durán[SUP] 2 [/SUP], Wagner José Fávaro[SUP] 2 [/SUP], Eder de Carvalho Pincinato[SUP] 2 [/SUP], Patricia Moriel[SUP] 5 [/SUP]
Affiliations
- PMID: 39500909
- PMCID: PMC11538513
- DOI: 10.1038/s41598-024-75740-3
When using the reverse-transcription quantitative polymerase chain reaction (RT-qPCR) technique for quantitative assessment of microRNA (miRNA) expression, normalizing data using a stable endogenous gene is essential; however, no universally adequate reference gene exists. Therefore, in this study, we aimed to determine, via the RNA-Seq technique, the most adequate endogenous normalizer for the expression assessment of plasma miRNAs in patients with coronavirus disease 2019 (COVID-19). Two massive sequencing procedures were performed (a) to identify differentially expressed miRNAs between patients with COVID-19 and healthy volunteers (n = 12), and (b) to identify differentially expressed miRNAs between patients with severe COVID-19 and those with mild COVID-19 (n = 8). The endogenous normalizer candidates were selected according to the following criteria: (1) the miRNA must have a fold regulation = 1; (2) the miRNA must have a p-value > 0.990; and (3) the miRNAs that were discovered the longest ago should be selected. Four miRNAs (hsa-miR-34a-3p, hsa-miR-194-3p, hsa-miR-17-3p, and hsa-miR-205-3p) met all criteria and were selected for validation by RT-qPCR in a cohort of 125 patients. Of these, only hsa-miR-205-3p was eligible endogenous normalizers in the context of COVID-19 because their expression was stable between the compared groups.
Keywords: COVID-19; Endogenous normalizer; MicroRNA; Reference gene.