Comparison of RNA Extraction Methods for Molecular Analysis of Oral Cytology

Author(s)

Alves, Monica Ghislaine Oliveira ^{[1, 2]} ; Perez-Sayans, Mario ^[3] ; Padin-Iruegas, Maria-Elena ^[4] ; Reboiras-Lopez, Maria Dolores ^[3] ; Suarez-Penaranda, Jose Manuel ^[5] ; Lopez-Lopez, Rafael ^[6] ; Carta, Celina Faig Lima ^[1] ; Issa, Jaqueline Scholz ^[7] ; Garcia-Garcia, Abel ^[3] ; Almeida, Janete Dias ^[1]

Affiliation

^[1] Univ Estadual Paulista, UNESP, Inst Sci & Technol, Dept Biosci & Oral Diag, Sao Jose Dos Campos, SP - Brazil

^[2] Univ Braz Cubas, Sch Dent, Mogi Das Cruzes - Brazil

^[3] Fac Med & Dent, Oral Med Oral Surg & Implantol Unit, Santiago De Compostela - Spain

^[4] Univ Vigo, Funct Biol & Hlth Sci Dept, Human Anat & Embriol Area, Pontevedra - Spain

^[5] Univ Hosp & Sch Med Santiago de Compostela, Dept Pathol, Forens Sci, La Coruna - Spain

^[6] Hosp Clin Univ Santiago de Compostela, Dept Med Oncol, La Coruna - Spain

^[7] Univ Sao Paulo, Smoking Cessat Program, Sch Med, Hosp Clin, Area Cardiol, Heart Inst, Sao Paulo - Brazil

• Document type: Journal article
• Source: ACTA STOMATOLOGICA CROATICA; v. 50, n. 2, p. 108-115, JUN 2016.
• Web of Science Citations: 1

Abstract

Objective of work: The aim of this study was to compare three methods of RNA extraction for molecular analysis of oral cytology to establish the best technique, considering its concentration and purity for molecular tests of oral lesions such as real-time reverse transcriptase reaction. Material and methods: The sample included exfoliative cytology from the oral cavity mucosa of patients with no visible clinical changes, using Orcellex Rovers Brush (R). The extraction of total RNA was performed using the following three techniques: 30 samples were extracted by Trizol (R) technique, 30 by the Directzol (TM) RNA Miniprep system and 30 by the RNeasy mini Kit. The absorbance was measured by spectrophotometer to estimate the purity. The estimated RNA concentration was obtained by multiplying the value of A260 (ng/mL) by 40. Statistical analysis of the obtained data was performed using GraphPad Prism 5.03 software with Student t, analysis of variance and Bonferroni tests, considering p <= 0.05. Results: Trizol (R) group revealed higher average concentration, followed by Direct-zol (TM) and Rneasy group. It was observed that the RNA Direct-zol (TM) group had the highest purity, followed by RNeasy and Trizol (R) groups, allowing for the two ratios. Conclusion: Considering all aspects, concentration, purity and time spent in the procedures, the Direct-zol (TM) group showed the best results. (AU)

• FAPESP's process: 12/05371-4 - Cytogenetic evaluation of smoking damage in oral mucosa
• Grantee: Janete Dias Almeida
• Support Opportunities: Regular Research Grants
• FAPESP's process: 13/06251-5 - Study of the effects caused by chronic smoking in oral mucosa cells
• Grantee: Mônica Ghislaine Oliveira Alves
• Support Opportunities: Scholarships in Brazil - Doctorate