Biorrepository saliva in genetic/molecular studies: evaluation of different DNA extraction protocols after long periods of storage

• Grant number: 14/24416-4
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: July 01, 2015
• End date: June 30, 2016
• Field of knowledge: Biological Sciences - Genetics - Human and Medical Genetics
• Principal Investigator: Lucimara Teixeira das Neves
• Grantee: Natália Ramos
• Host Institution: Faculdade de Odontologia de Bauru (FOB). Universidade de São Paulo (USP). Bauru , SP, Brazil

Abstract

In the field of genetic and molecular analysis it is fundamentally important for results to effectively use quality biological materials. Among the biological materials where DNA can be isolated, blood is by far the most widely used, yet its collection method is invasive; additionally a collector needs specialized training for collection. Therefore the aim of this study was to analyze the quality and quantity of genomic DNA extracted from saliva samples, a method gaining recognition as an alternative source for DNA isolation. Besides being noninvasive in terms of collection and thereby tolerated by most individuals, saliva collection reduces the risk of contamination when compared to other methods. Unlike blood samples, saliva samples can be frozen for long periods of time, enabling the implementation of biobanks for such biological samples. The objectives of this study, in particular, is to analyze the quality, integrity and purity of genomic DNA, using four different extraction protocols; three of these protocols are from commercially available kits. After extraction, genomic DNA will be submitted to spectrophotometric analysis, sequencing and genotyping of various sequences including the following: exon 3 for the IRF6 gene, polymorphism rs 12532, and the MSX1 gene. These analyses should verify the influence of freezing saliva samples at -20°C for a period of 2 to 6 years, and determine the best extraction protocol used for molecular experiments performed (sequencing and genotyping by real time PCR).