| Autor: |
Yui Mizumoto-Teramura, Teru Kamogashira, Kenji Kondo, Tatsuya Yamasoba |
| Jazyk: |
angličtina |
| Rok vydání: |
2024 |
| Předmět: |
|
| Zdroj: |
MethodsX, Vol 13, Iss , Pp 102818- (2024) |
| Druh dokumentu: |
article |
| ISSN: |
2215-0161 |
| DOI: |
10.1016/j.mex.2024.102818 |
| Popis: |
Real-time PCR with intercalating dyes can only be performed once. The expensive fluorescent hydrolysis probes are target specific and are suitable to detect multiplex targets. Uracil-DNA N-glycosylase (UNG), which specifically hydrolyzes and degrades any uracil-containing PCR products, is often applied before PCR to reduce carryover contamination. We developed an optimized protocol for recovering DNA from PCR products and revaluating by real-time PCR with intercalating dye using UNG processing, which is particularly useful when the sample volume is very small and insufficient for multiple assays of real-time PCR. • A real-time PCR master mix with dUTP instead of dTTP was used. • UNG at 1 % and 10 % concentrations of PCR product volumes were used for the first and second processing. • The second real-time PCR was performed with different primer pairs than the first real-time PCR. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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