Журнал биопереработки и биотехники

A New Procedure for DNA Isolation from Saliva Samples and Comparative Analysis of Quality Indicators

Abstract

Carrillo Avila JA, De la Puente R, Pinaglia Tobaruela G, Aguilar Quesada R, Rueda Medina B, Correa Rodriguez M and Espin Vallejo L

Background: Large population-based studies involving thousands of participants are needed for research on genetic diseases and epidemiologic studies. Saliva samples are a non-invasive and efficient DNA source for mass collection. The establishment of new optimized DNA isolation procedures from saliva and the determination of the most effective quality indicator are essential for this purpose.

Methods: DNA was extracted from 112 saliva samples utilizing a novel method. Samples were pre-treated with Protease for 1 h at 56°C, and reagents from a kit for blood samples were used in the Chemagic MSM-I automated instrument with a specifically designed saliva protocol for the Chemagic software. DNA quality was estimated by spectrophotometry, fluorometry, qPCR, SPUD assay and the Agilent 2200 Tape Station.

Results: An average DNA yield of 52.58 ± 33.77 μg was obtained with no significant differences between males and females. A₂₆₀/A₂₈₀ and A₂₆₀/A₂₃₀ ratios of 1.84 ± 0.123 and 1.56 ± 0.297 were obtained respectively. A DIN value of 6.83 ± 0.90 was observed with a satisfactory functionality calculated by qPCR analysis. On the other hand, significant differences were observed between spectrophotometry, fluorometry and qPCR quantification methods in spite of the low amount of contaminants detected.

Conclusion: Collecting as many samples as possible is necessary to establish DNA cohorts that represent the whole population. The non-invasive procedure described in this work guarantees a large amount of DNA from saliva samples valid for any downstream molecular applications, with an important reduction in costs. Additionally, an innovative comparison between the DIN values and conventional DNA quality indicators is shown.