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Pregled bibliografske jedinice broj: 1098598

Pre-amplification as a method for improvement of quantitative RT-PCR analysis of circulating miRNAs


Sekovanić, Ankica; Dorotić, Adrijana; Jurasović, Jasna; Pašalić, Daria; Kovačić, Jelena; Stasenko, Sandra; Mioč, Tatjana; Piasek, Martina; Orct, Tatjana
Pre-amplification as a method for improvement of quantitative RT-PCR analysis of circulating miRNAs // Biochemia medica, 31 (2021), 1; 010901, 6 doi:10.11613/BM.2021.010901 (međunarodna recenzija, članak, znanstveni)


CROSBI ID: 1098598 Za ispravke kontaktirajte CROSBI podršku putem web obrasca

Naslov
Pre-amplification as a method for improvement of quantitative RT-PCR analysis of circulating miRNAs

Autori
Sekovanić, Ankica ; Dorotić, Adrijana ; Jurasović, Jasna ; Pašalić, Daria ; Kovačić, Jelena ; Stasenko, Sandra ; Mioč, Tatjana ; Piasek, Martina ; Orct, Tatjana

Izvornik
Biochemia medica (1330-0962) 31 (2021), 1; 010901, 6

Vrsta, podvrsta i kategorija rada
Radovi u časopisima, članak, znanstveni

Ključne riječi
blood plasma ; circulating microRNAs ; epigenetics ; pre-amplification ; RT-PCR

Sažetak
Introduction: The assessment of circulating miRNAs is challenging and still limited due to their low concentrations, small size and lack of reference values in human biological samples. Pre-amplification of complementary DNAs may facilitate reliable miRNA quantification. The aim of our study was to evaluate the efficacy of pre- amplification as a step to increase the sensitivity of qPCR analysis for five candidate circulating miRNAs presumably related to toxic metals and cigarette smoke exposure: miR-1537, miR-190b, miR-16, miR-21, and miR-146a. Materials and methods: Candidate miRNAs expression was analysed in plasma samples of 19 mother-newborn pairs. For isolation, transcription, pre- amplification and qPCR quantification kits and protocols by Qiagen (Hilden, Germany) were used. Paired t-test or Wilcoxon rank test were used to compare miRNAs expression levels with and without a pre- amplification step prior to qPCR, separately in maternal and cord plasma. Intraclass correlation (ICC) was calculated as an agreement measure between procedures for each miRNA. Results: Pre-amplification facilitated the detection of all assayed miRNAs with an overall cycle threshold (CT) improvement of 6.6 ± 0.89 (P < 0.05). Excellent ICCs (> 0.90) were found between data for preamplified and not preamplified miR-16, miR- 21 and miR-146a. However, these correlations for low expressed miR-190b were moderate (0.79 in maternal ; 0.61 in cord plasma) and poor for miR-1537 (0.49 in maternal ; no correlation in cord plasma). Conclusion: Pre-amplification is a useful, necessary step in the analysis of miR-1537 and miR-190b as a reliable procedure facilitating extracellular miRNA expression detection in human plasma by real-time PCR quantification.

Izvorni jezik
Engleski

Znanstvena područja
Temeljne medicinske znanosti, Kliničke medicinske znanosti, Javno zdravstvo i zdravstvena zaštita

Napomena
Submitted: May 29, 2020
Accepted: Sep 28, 2020



POVEZANOST RADA


Projekti:
IP-2016-06-1998 - Procjena svakodnevne izloženosti metalima i osobne osjetljivosti majke kao čimbenika razvojnoga podrijetla zdravlja i bolesti (METALORIGINS) (Piasek, Martina, HRZZ - 2016-06) ( POIROT)

Ustanove:
Institut za medicinska istraživanja i medicinu rada, Zagreb,
Klinička bolnica "Merkur",
Medicinski fakultet, Zagreb,
Klinička bolnica "Sveti Duh"

Citiraj ovu publikaciju

Sekovanić, Ankica; Dorotić, Adrijana; Jurasović, Jasna; Pašalić, Daria; Kovačić, Jelena; Stasenko, Sandra; Mioč, Tatjana; Piasek, Martina; Orct, Tatjana
Pre-amplification as a method for improvement of quantitative RT-PCR analysis of circulating miRNAs // Biochemia medica, 31 (2021), 1; 010901, 6 doi:10.11613/BM.2021.010901 (međunarodna recenzija, članak, znanstveni)
Sekovanić, A., Dorotić, A., Jurasović, J., Pašalić, D., Kovačić, J., Stasenko, S., Mioč, T., Piasek, M. & Orct, T. (2021) Pre-amplification as a method for improvement of quantitative RT-PCR analysis of circulating miRNAs. Biochemia medica, 31 (1), 010901, 6 doi:10.11613/BM.2021.010901.
@article{article, year = {2021}, pages = {6}, DOI = {10.11613/BM.2021.010901}, chapter = {010901}, keywords = {blood plasma, circulating microRNAs, epigenetics, pre-amplification, RT-PCR}, journal = {Biochemia medica}, doi = {10.11613/BM.2021.010901}, volume = {31}, number = {1}, issn = {1330-0962}, title = {Pre-amplification as a method for improvement of quantitative RT-PCR analysis of circulating miRNAs}, keyword = {blood plasma, circulating microRNAs, epigenetics, pre-amplification, RT-PCR}, chapternumber = {010901} }
@article{article, year = {2021}, pages = {6}, DOI = {10.11613/BM.2021.010901}, chapter = {010901}, keywords = {blood plasma, circulating microRNAs, epigenetics, pre-amplification, RT-PCR}, journal = {Biochemia medica}, doi = {10.11613/BM.2021.010901}, volume = {31}, number = {1}, issn = {1330-0962}, title = {Pre-amplification as a method for improvement of quantitative RT-PCR analysis of circulating miRNAs}, keyword = {blood plasma, circulating microRNAs, epigenetics, pre-amplification, RT-PCR}, chapternumber = {010901} }

Časopis indeksira:


  • Current Contents Connect (CCC)
  • Web of Science Core Collection (WoSCC)
    • Science Citation Index Expanded (SCI-EXP)
    • SCI-EXP, SSCI i/ili A&HCI
  • Scopus
  • MEDLINE


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