Comparative Analysis of RNA Extraction Quality Using TRIzol-Based and Non-TRIzol Methods in Endometriosis Lesion Tissue

Ocktariyana Ocktariyana; Bertha Octarina; Shungai Mutiara Aini; Refai Refai; Irsan Saleh; Fatimah Usman; Adnan Abadi; Asmarinah Asmarinah

doi:10.31964/mltj.v12i1.744

Authors

Ocktariyana Ocktariyana Department of Midwifery, Poltekkes Kemenkes Palembang, Palembang, Indonesia
Bertha Octarina Department of Obstetrics and Gynecology, Faculty of Medicine Universitas Sriwijaya, Dr. Mohammad Hoesin General Hospital, Palembang, Indonesia
Shungai Mutiara Aini Medical Laboratory Technology Faculty, Poltekkes Kemenkes Palembang, Palembang, Indonesia
Refai Refai Medical Laboratory Technology Faculty, Poltekkes Kemenkes Palembang, Palembang, Indonesia
Irsan Saleh Department of Pharmacology, Faculty of Medicine Universitas Sriwijaya, Palembang, Indonesia
Fatimah Usman Department of Obstetrics and Gynecology, Faculty of Medicine Universitas Sriwijaya, Dr. Mohammad Hoesin General Hospital, Palembang, Indonesia
Adnan Abadi Department of Obstetrics and Gynecology, Faculty of Medicine Universitas Sriwijaya, Dr. Mohammad Hoesin General Hospital, Palembang, Indonesia
Asmarinah Asmarinah Department of Biology Medicine, Faculty of Medicine Universitas Indonesia, Jakarta, Indonesia

DOI:

https://doi.org/10.31964/mltj.v12i1.744

Keywords:

Endometriosis, RNA extraction, RNA quality, tissue homogenization, TRIzol

Abstract

Ribonucleic acid (RNA) is a crucial molecule in gene expression analysis, yet its isolation from solid tissues, such as endometriotic lesions, poses significant challenges due to complex tissue structure and high RNase activity. TRIzol is a widely utilized reagent for RNA extraction, known for its efficacy in lysing cells and stabilizing RNA. This study aimed to compare the quality of RNA extracts from endometriotic lesion tissue samples using TRIzol-based and non-TRIzol-based methods. A quasi-experimental design was employed, involving a control group and an experimental group, with non-randomized sample allocation. A total of 28 samples were analyzed in both the control and treatment groups. The results demonstrated that TRIzol treatment significantly increased the median RNA concentration from endometriotic lesion tissues, yielding 288.6 ng/μL compared to 118.2 ng/μL in the non-TRIzol group (p=0.001). Furthermore, there was a significant difference between TRIzol and non-TRIzol groups in RNA purity based on the Å260/280 ratio (median: 2.00; p-value:0.024). However, there wasn’t a significant difference in the Å260/230 ratio (median: 2.11; p-value: 0.247). In conclusion, these findings indicate that while TRIzol extraction provides higher RNA yield, the non‑TRIzol method ensures slightly superior sample purity, supporting its reliability for downstream transcriptomic analyses.

References

Becker, C. M., Bokor, A., Heikinheimo, O., Horne, A., Jansen, F., Kiesel, L., King, K., Kvaskoff, M., Nap, A., & Petersen, K. (2022). ESHRE guideline: endometriosis. Human Reproduction Open, 2022(2), hoac009. https://doi.org/10.1093/ hropen/hoac009.

Bhat, T. A., & Hakeem, K. R. (2023). Biotechnologies and Genetics in Plant Mutation Breeding. Biotechnologies and Genetics in Plant Mutation Breeding, 2, 1–272. https://doi.org/10.1201/9781003305088

Boom, R., Sol, C. J., Salimans, M. M., Jansen, C. L., Wertheim-van Dillen, P. M., & van der Noordaa, J. (1990). Rapid and simple method for purification of nucleic acids. Journal of Clinical Microbiology, 28(3), 495–503. https://doi.org/10.1128/JCM.28.3.495-503.1990

Bulun, S. E. (2009). Endometriosis. The New England Journal of Medicine, 360(3), 268–279. https://doi.org/10.1056/NEJMra0804690

Chen, X., Cui, D., & Chen, J. (2012). Microfluidic Chips for DNA Extraction and Purification. In On-Chip Pretreatment of Whole Blood by Using MEMS Technology (pp. 84–109). Bentham Science Publishers. https://doi.org/ 10.2174/978160805147211201010084.

Febriyani, P. D. Z., Syailendra, A., Dani, H., Ocktariyana, Pasha, A. R., & Aini, S. M. (2026). Evaluation of tris–EDTA pre-hydrated non-FTA dried blood spot as a low-cost medium for room-temperature DNA sample transport. Jurnal Teknologi Laboratorium, 15(1), 48–58. https://doi.org/10.29238/TEKNOLABJOURNAL. V15I1.712

Fisher Scientific, T. (2025). Procedural guidelines TRIzolTM Reagent. https://www.thermofisher.com/trizolfaqs. Accessed on June 6, 2026.

Gallagher, S. R. (2011). Quantitation of DNA and RNA with absorption and fluorescence spectroscopy. Current Protocols in Molecular Biology, 93(SUPPL.93), A.3D.1-A.3D.14. https://doi.org/10.1002/0471142727. MBA03DS93;SUBPAGE:STRING:ABSTRACT;REQUESTEDJOURNAL:JOURNAL:19343647;ISSUE:ISSUE:DOI. Accessed on June, 6 2026.

Gandhi, V., O’Brien, M. H., & Yadav, S. (2020). High-quality and high-yield RNA extraction method from whole human saliva. Journals.Sagepub.ComV Gandhi, MH O’Brien, S YadavBiomarker Insights, 2020•journals.Sagepub.Com, 15. https://doi.org/10.1177/1177271920929705

Gautam, A. (2022). DNA and RNA Isolation Techniques for Non-experts. https://link.springer.com/content/pdf/10.1007/978-3-030-94230-4.pdf. Accessed on June, 6 2026.

Hasegawa, T., Hapsari, D., & Iwahashi, H. (2021). Challenges for Accurate Quantification of RNA. Reviews in Agricultural Science, 9(0), 1–19. https://doi.org/10.7831/ras.9.0_1

Haupenthal, J., Baehr, C., Kiermayer, S., Zeuzem, S., & Piiper, A. (2006). Inhibition of RNAse A family enzymes prevents degradation and loss of silencing activity of siRNAs in serum. Biochemical Pharmacology, 71(5), 702-710. https://doi.org/10.1016/j.bcp.2005.11.015.

Hongjaisee, S., Jabjainai, Y., Sakset, S., Preechasuth, K., Ngo-Giang-Huong, N., & Khamduang, W. (2022). Comparison of simple RNA extraction methods for molecular diagnosis of hepatitis C virus in plasma. Diagnostics, 12(7), 1599. https://doi.org/10.3390/diagnostics12071599

Hwan Shin, J. (2018). Nucleic Acid Extraction and Enrichment. Advanced Techniques in Diagnostic Microbiology: Volume 1: Techniques, Third Edition, 273–292. https://doi.org/10.1007/978-3-319-33900-9_13/TABLES/3.

Imanaka, S., Maruyama, S., Kimura, M., Nagayasu, M., & Kobayashi, H. (2020). Towards an understanding of the molecular mechanisms of endometriosis-associated symptoms. World Academy of Sciences Journal, 2(4), 12. https://doi.org/10.3892/wasj.2020.53.

Laganà, A. S., Garzon, S., Götte, M., Viganò, P., Franchi, M., Ghezzi, F., & Martin, D. C. (2019). The pathogenesis of endometriosis: molecular and cell biology insights. International Journal of Molecular Sciences, 20(22), 5615. https://doi.org/10.3390/ijms20225615.

Lakhotia, Subhash & Ranganath, H.. (2024). Experiments with Drosophila for Biology Courses. 10.5281/zenodo.11206949.

Liu, H., Luo, M., & Wen, J. (2014). mRNA stability in the nucleus. Journal of Zhejiang University SCIENCE B, 15(5), 444–454. https://doi.org/10.1631/jzus.B1400088.

Martins, R., Queiroz, J. A., & Sousa, F. (2014). Ribonucleic acid purification. Journal of Chromatography A, 1355, 1–14. https://doi.org/10.1016/ J.CHROMA.2014.05.075.

Masoomi-Aladizgeh, F., Jabbari, L., Khayam Nekouei, R., Aalami, A., Atwell, B. J., & Haynes, P. A. (2023). A universal protocol for high-quality DNA and RNA isolation from diverse plant species. Journals.Plos.Org, 18(12 December). https://doi.org/10.1371/JOURNAL.PONE.0295852

McDowall, K. J. (2007). RNA Stability: Chemistry, Measurement and Modulation. Wiley Encyclopedia of Chemical Biology, 1–13. https://doi.org/10.1002/9780470048672.wecb521.

Mohanty, S., Singh, M., Jena, S., Kumar, S., & Jha, S. (2025). RNA adsorption onto iron oxide nanoparticle have stability from nuclease. International Journal of Biological Macromolecules, 322(Pt 2), 146655. https://doi.org/10.1016/j.ijbiomac.2025.146655.

Nnoaham, K. E., Hummelshoj, L., Webster, P., D’Hooghe, T., De Cicco Nardone, F., De Cicco Nardone, C., Jenkinson, C., Kennedy, S. H., & Zondervan, K. T. (2011). Impact of endometriosis on quality of life and work productivity: A multicenter study across ten countries. Fertility and Sterility, 96(2). https://doi.org/10.1016/j.fertnstert.2011.05.090

Non-Experts, A. G.-D. and R. I. T. for, & 2022, undefined. (2022). RNA isolation by the Guanidinium-acid-phenol method. Springer, 41–46. https://doi.org/10.1007/978-3-030-94230-4_4

Nouvel, A., Laget, J., Duranton, F., Leroy, J., Desmetz, C., Servais, M. D., de Préville, N., Galtier, F., Nocca, D., Builles, N., Rebuffat, S., & Lajoix, A. D. (2021). Optimization of RNA extraction methods from human metabolic tissue samples of the COMET biobank. Scientific Reports 2021 11:1, 11(1), 20975-. https://doi.org/10.1038/s41598-021-00355-x

Nouvel, A., Laget, J., Duranton, F., Leroy, J., Desmetz, C., Servais, M.-D., De Préville, N., Galtier, F., Nocca, D., Builles, N., Rebuffat, S., & Lajoix, A.-D. (123 C.E.). Optimization of RNA extraction methods from human metabolic tissue samples of the COMET biobank. Nature.ComA Nouvel, J Laget, F Duranton, J Leroy, C Desmetz, MD Servais, N de Préville, F GaltierScientific Reports, 2021•nature.Com, 11, 20975. https://doi.org/10.1038/s41598-021-00355-x

Nur, A., & Yamamoto, Z. (2022). Saliva as a source of human genomic DNA. Jurnal Kedokteran Syiah Kuala, 22(2). https://jurnal.usk.ac.id/JKS/article/viewFile/ 23386/17108.

Obino, D., Vassalli, M., Franceschi, A., Alessandrini, A., Facci, P., & Viti, F. (2021). An overview on microfluidic systems for nucleic acids extraction from human raw samples. Sensors, 21(9), 3058. https://doi.org/10.3390/s21093058

Ocktariyana. (2020). Pain-Endometriosis In Molecular Perspective (1st ed., Vol. 1). Chakra Brahmanda Lentera. https://www.researchgate.net/profile/Ocktariyana-Ocktariyana/publication/357518032_Nyeri_Pada_Endometriosis_Dalam_Perspektif_Molekuler/links/61d23cfdb6b5667157c2e96f/Nyeri-Pada-Endometriosis-Dalam-Perspektif-Molekuler.pdf. Accessed on June 6, 2026.

Ocktariyana, O., Lestari, D., & Asmarinah, A. (2022). Analisis Kemurnian Ekstraksi Asam Ribonukleat Pada Sampel Darah Menstruasi. JPP (Jurnal Kesehatan Poltekkes Palembang), 17(2), 238–242. https://doi.org/ 10.36086/jpp.v17i2.

Pagani, S., Maglio, M., Sicuro, L., Fini, M., Giavaresi, G., & Brogini, S. (2023). RNA Extraction from Cartilage: Issues, Methods, Tips. International Journal of Molecular Sciences, 24(3), 2120. https://doi.org/10.3390/IJMS24032120

Rolfs, F., de Goeij-de Haas, R. R., Knol, J. C., Piersma, S. R., & Jimenez, C. R. (2023). Phosphoproteomics After Guanidinium Thiocyanate Extraction of Tissue Biopsies. Springer, 2718, 285–302. https://doi.org/10.1007/978-1-0716-3457-8_16

Roy, D., Tomo, S., Modi, A., Purohit, P., MethodsX, P. S.-, & 2020, undefined. (2020). Optimising total RNA quality and quantity by phenol-chloroform extraction method from human visceral adipose tissue: A standardisation study. Elsevier, 2134, 41–51. https://doi.org/10.1007/978-1-0716-0459-5_5

Suwartono, H., Kaput, J. A., Astijani, N. M., & Saranga, D. (2022). Endometriosis Subkutan: Laporan Kasus Subcutaneous Endometriosis: Case Report. Jurnal Medical Profession (Medpro), 4(1), 29–35. https://jurnal.fk.untad.ac.id/index.php/medpro/article/view/585

Tan, S. C., & Yiap, B. C. (2009). DNA, RNA, and protein extraction: The past and the present. Journal of Biomedicine and Biotechnology, 2009. https://doi.org/10.1155/2009/574398

Taylor, H. S., Adamson, G. D., Diamond, M. P., Goldstein, S. R., Horne, A. W., Missmer, S. A., Snabes, M. C., Surrey, E., & Taylor, R. N. (2018). An evidence-based approach to assessing surgical versus clinical diagnosis of symptomatic endometriosis. International Journal of Gynecology & Obstetrics, 142(2), 131–142. https://doi.org/10.1002/ijgo.12521

Taylor, H. S., Giudice, L. C., Lessey, B. A., Abrao, M. S., Kotarski, J., Archer, D. F., Diamond, M. P., Surrey, E., Johnson, N. P., Watts, N. B., Gallagher, J. C., Simon, J. A., Carr, B. R., Dmowski, W. P., Leyland, N., Rowan, J. P., Duan, W. R., Ng, J., Schwefel, B., Thomas, J. W., … Chwalisz, K. (2017). Treatment of endometriosis-associated pain with elagolix, an oral GnRH antagonist. New England Journal of Medicine, 377(1), 28–40. https://doi.org/10.1056/NEJMoa1700089

Widen, R. H., & Silbert, S. (2016). Nucleic Acid Extraction in Diagnostic Virology. Clinical Virology Manual, 117–128. https://doi.org/10.1128/9781555819156. ch10Digital Object Identifier (DOI)

Yin, M., Zhai, L., Wang, J., Yu, Q., Li, T., Xu, X., Guo, X., Mao, X., Zhou, J., & Zhang, X. (2022). Comprehensive analysis of RNA-seq in endometriosis reveals competing endogenous RNA network composed of circRNA, lncRNA and mRNA. Frontiers in Genetics, 13, 828238. https://doi.org/10.3389/fgene.2022.828238

Zhang, H., Liu, Y., Yu, B., & Lu, R. (2023). An Optimized TRIzol-Based Method for Isolating RNA from Adipose Tissue. BioTechniques, 74(5), 203–209. https://doi.org/10.2144/BTN-2022-0120.