Development of a low-cost and robust SYBR Green buffer system for qPCR

• Grant number: 17/03263-3
• Support Opportunities: Regular Research Grants
• Start date: July 01, 2017
• End date: June 30, 2019
• Field of knowledge: Health Sciences - Dentistry - Endodontics
• Principal Investigator: Rafael Nobrega Stipp
• Grantee: Rafael Nobrega Stipp
• Host Institution: Faculdade de Odontologia de Piracicaba (FOP). Universidade Estadual de Campinas (UNICAMP). Piracicaba , SP, Brazil

Abstract

Aim: Develop a low-cost reaction mixture for Quantitative PCR (qPCR) with specificity, efficiency, and sensitivity equivalent to the foreign commercial kits. Methods: Mixtures will be with varied concentrations of SYBR® Green, MgCl2 and Platinum Taq DNA Polymerase Brazil (core reagents). The additives bovine seric albumin, dimethyl sulfoxide, formamide, polyethylene glycol 400, polyoxyethylene-octyl-phenyl-ether, trehalose and polysorbate-20 willl be added alone or combined and evaluated as reactions enhancers. During the developing process, reactions will be conducted with Porphyromonas gingivalis genomic DNA (300 to 0.0003 ng) and specie-specific primers. To ensure the applicability, the blends with the best performances will be assayed against a range of DNA from other species and its corresponding primers. CT (Cycle threshold) values and melting curves will be used to compare mixtures to the commercial Power SYBR Green qPCR Master Mix Kit©. The current price of a qPCR reaction can be lowered at 10% by using non industrialized mixture. (AU)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

LEI, L.; STIPP, R. N.; CHEN, T.; WU, S. Z.; HU, T.; DUNCAN, M. J.. Activity of Streptococcus mutans VicR Is Modulated by Antisense RNA. JOURNAL OF DENTAL RESEARCH, v. 97, n. 13, p. 1477-1484, DEC 2018. (17/03263-3)

SALES, LUCIANA SOLERA; GUIMARAES, GUSTAVO NARVAES; WIJESINGHE, GAYAN KANCHANA; SILVA MOREIRA, KELLY MARIA; JOIA, FELIPE; STIPP, RAFAEL NOBREGA; AZEVEDO RODRIGUES, LIDIANY KARLA; NOBRE-DOS-SANTOS, MARINES; STEINER-OLIVEIRA, CAROLINA. Addition of hydrogen peroxide to methylene blue conjugated to beta-cyclodextrin in photodynamic antimicrobial chemotherapy in S. mutans biofilm. Photodiagnosis and Photodynamic Therapy, v. 28, p. 226-233, DEC 2019. (17/03263-3)