Preservation of milk in liquid nitrogen during sample collection does not affect the RNA quality for RNA-seq analysis
| dc.contributor.author | Jiménez Montenegro, Lucía | |
| dc.contributor.author | Alfonso Ruiz, Leopoldo | |
| dc.contributor.author | Soret Lafraya, Beatriz | |
| dc.contributor.author | Mendizábal Aizpuru, José Antonio | |
| dc.contributor.author | Urrutia Vera, Olaia | |
| dc.contributor.department | Agronomía, Biotecnología y Alimentación | es_ES |
| dc.contributor.department | Agronomia, Bioteknologia eta Elikadura | eu |
| dc.contributor.department | Institute on Innovation and Sustainable Development in Food Chain - ISFOOD | en |
| dc.contributor.funder | Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa | |
| dc.contributor.funder | Gobierno de Navarra / Nafarroako Gobernua | |
| dc.date.accessioned | 2025-07-02T14:41:06Z | |
| dc.date.available | 2025-07-02T14:41:06Z | |
| dc.date.issued | 2025-05-24 | |
| dc.date.updated | 2025-07-02T14:22:15Z | |
| dc.description.abstract | Background. Standard procedures for milk sample collection for transcriptome analysis use ice as preservation method, which can afect the RNA stability and requires immediate sample processing. These problems would be eased if the milk samples could be snap-frozen in liquid nitrogen. This study describes the applicability of a new method for milk sample collection and subsequent RNA extraction from milk fat globules, determining whether the quality, integrity and quantity of the RNA extracts met the minimum requirements for downstream RNA-seq. Results. The quality of the extracts measured by A260/280 ratio and the Integrity and Quality (IQ) values obtained fulflled the reference values of 1.9 - 2.1 (P10.05) and failed to meet the RIN≥7 benchmark for RNA-seq (P>0.05). Milk fat globules contain low molecular-weight RNA fragments and minimal 18S and 28S rRNA, suggesting low RIN values were inherent to sample type. Likewise, the RNA concentration from milk fat globules were generally low (120.43±22.27 ng/µL, 102.87±15.64 ng/µL and 109.43±22.69 ng/µL, measured by Nanodrop, Qubit HS and QuanTI Ribogreen, respectively). Nevertheless, RNA-seq yielded 52.7 million paired-end reads per sample. The raw reads passed all quality control parameters having the same sequence-read lengths (151 bp), 100% base-coverage, 49% GC base content, and base quality scores of 36, enabling successful transcriptome profling. Moreover, milk proteins were identifed as the most abundant transcripts in MFG in the analysis of the most expressed genes, indicating that the sequenced reads would accurately refect the transcriptome of this milk fraction. Conclusions. Milk preservation in liquid nitrogen is a suitable sample collection method that overcomes the limitations of immediate sample processing required if ice is used. Thus, this procedure, together with the subsequent RNA isolation from milk fat globules and its sequencing by RNA-seq, would provide a practical and a non-invasive method for measuring the mammary epithelial cell transcriptome, improving the feasibility of conducting studies related to mammary gland and lactation physiology. | en |
| dc.description.sponsorship | Open Access funding provided by Universidad Pública de Navarra. This research was funded by the Public University of Navarre within the framework of Young Researcher Projects 2022 and by the Department of University, Innovation, and Digital Transformation of the Government of Navarre, grant number 0011-1408-2022-000024 (RES 95E/2022 of May 17). | |
| dc.format.mimetype | application/pdf | |
| dc.identifier.citation | Jiménez-Montenegro, L., Alfonso, L., Soret, B., Mendizabal, J. A., Urrutia, O. (2025) Preservation of milk in liquid nitrogen during sample collection does not affect the RNA quality for RNA-seq analysis. BMC Genomics, 26, 1-14. https://doi.org/10.1186/s12864-025-11707-6. | |
| dc.identifier.doi | 10.1186/s12864-025-11707-6 | |
| dc.identifier.issn | 1471-2164 | |
| dc.identifier.uri | https://academica-e.unavarra.es/handle/2454/54372 | |
| dc.language.iso | eng | |
| dc.publisher | BMC | |
| dc.relation.ispartof | BMC Genomics 26, 525, (2025) | |
| dc.relation.projectID | info:eu-repo/grantAgreement/Gobierno de Navarra//0011-1408-2022-000024/ | |
| dc.relation.publisherversion | https://doi.org/10.1186/s12864-025-11707-6 | |
| dc.rights | © The Author(s) 2025. Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License, which permits any non-commercial use, sharing, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if you modified the licensed material. | |
| dc.rights.accessRights | info:eu-repo/semantics/openAccess | |
| dc.rights.uri | http://creativecommons.org/licenses/by-nc-nd/4.0/ | |
| dc.subject | Liquid nitrogen | en |
| dc.subject | Mammary gland | en |
| dc.subject | Milk | en |
| dc.subject | Milk fat globule | en |
| dc.subject | RNA isolation | en |
| dc.subject | RNA-sequencing | en |
| dc.title | Preservation of milk in liquid nitrogen during sample collection does not affect the RNA quality for RNA-seq analysis | en |
| dc.type | info:eu-repo/semantics/article | |
| dc.type.version | info:eu-repo/semantics/publishedVersion | |
| dspace.entity.type | Publication | |
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