Open Access

Comparison and optimisation of microRNA extraction from the plasma of healthy pregnant women

  • Authors:
    • Victoria L. Parker
    • Bryony F. Cushen
    • Eleftherios Gavriil
    • Benjamin Marshall
    • Sarah Waite
    • Allan Pacey
    • Paul R. Heath
  • View Affiliations

  • Published online on: February 5, 2021     https://doi.org/10.3892/mmr.2021.11897
  • Article Number: 258
  • Copyright: © Parker et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Circulating microRNA (miRNA) biomarkers are implicated in the diagnosis, monitoring and prediction of various disease processes. Before embarking upon biomarker discovery, miRNA extraction techniques must first be optimised in the biofluid and population under study. Using plasma from a healthy pregnant woman, it was attempted to optimise and compare the performance of two commercially available miRNA extraction kits; Qiagen (miRNeasy Serum/Plasma) and Promega (Maxwell® RSC miRNA from Tissue or Plasma or Serum). Sample miRNA content (concentration and percentage) was assessed using Agilent Bioanalyzer Small RNA chips and reverse transcription‑quantitative PCR (RT‑qPCR) using four constitutively expressed miRNAs (hsa‑miR‑222‑3p, hsa‑let‑7i‑3p, hsa‑miR‑148‑3p and hsa‑miR‑30e‑5p). Quality control spike‑ins monitored RNA extraction (UniSp2, 4 and 5) and cDNA synthesis (UniSp6, cel‑miR‑39‑3p) efficiency. Optimisation approaches included: i) Starting volume of plasma; the addition of ii) Proteinase K; iii) a RNA bacteriophage carrier (MS2); and iv) a glycogen carrier. The two kits exhibited equivalence in terms of miRNA recovery based on Bioanalyzer and RT‑qPCR ΔΔCq results. Optimisation attempts for both kits failed to improve upon miRNA content compared with standard methodology. Comparing the standard methodology, the Qiagen kit was more consistent (smaller variance of ΔCq values) compared with the Promega kit. The standard methodology of either kit would be suitable for the investigation of miRNA biomarkers in a healthy pregnant population.
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April-2021
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Spandidos Publications style
Parker VL, Cushen BF, Gavriil E, Marshall B, Waite S, Pacey A and Heath PR: Comparison and optimisation of microRNA extraction from the plasma of healthy pregnant women. Mol Med Rep 23: 258, 2021.
APA
Parker, V.L., Cushen, B.F., Gavriil, E., Marshall, B., Waite, S., Pacey, A., & Heath, P.R. (2021). Comparison and optimisation of microRNA extraction from the plasma of healthy pregnant women. Molecular Medicine Reports, 23, 258. https://doi.org/10.3892/mmr.2021.11897
MLA
Parker, V. L., Cushen, B. F., Gavriil, E., Marshall, B., Waite, S., Pacey, A., Heath, P. R."Comparison and optimisation of microRNA extraction from the plasma of healthy pregnant women". Molecular Medicine Reports 23.4 (2021): 258.
Chicago
Parker, V. L., Cushen, B. F., Gavriil, E., Marshall, B., Waite, S., Pacey, A., Heath, P. R."Comparison and optimisation of microRNA extraction from the plasma of healthy pregnant women". Molecular Medicine Reports 23, no. 4 (2021): 258. https://doi.org/10.3892/mmr.2021.11897