Development of Plasmid Calibrators for Absolute Quantification of the β-parvalbumin gene in Lophius piscatorius
The result's identifiers
Result code in IS VaVaI
<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00027022%3A_____%2F23%3AN0000005" target="_blank" >RIV/00027022:_____/23:N0000005 - isvavai.cz</a>
Alternative codes found
RIV/00216208:11310/23:10469637 RIV/60461373:22330/23:43927681
Result on the web
<a href="https://link.springer.com/article/10.1007/s00217-023-04357-z" target="_blank" >https://link.springer.com/article/10.1007/s00217-023-04357-z</a>
DOI - Digital Object Identifier
<a href="http://dx.doi.org/10.1007/s00217-023-04357-z" target="_blank" >10.1007/s00217-023-04357-z</a>
Alternative languages
Result language
angličtina
Original language name
Development of Plasmid Calibrators for Absolute Quantification of the β-parvalbumin gene in Lophius piscatorius
Original language description
The real-time quantitative PCR (qPCR) calibration curves are highly reproducible and allow the generation of specific, sensitive, and reproducible data that can be used for gene quantification. However, it is important to rigorously validate the external calibration curve model in qPCR since absolute quantification is dependent on the standards used. We present a method for standardising qPCR-based quantification of the β-parvalbumin (β-pvalb) gene of Lophius piscatorius, a major fish allergen, using a plasmid DNA (pDNA) calibrator. In parallel experiments, standard curves were generated and compared from the genomic DNA (gDNA) isolated from L. piscatorius and pDNA carrying the target, pvalb. The commutability of pDNA and gDNA calibrators for the quantification of β-pvalb was assessed by employing a TaqMan qPCR, targeting the second intron of the pvalb gene of L. piscatorius. Higher PCR efficiencies, good linearity, and lower standard deviation (S.D.) values were observed with pDNA instead of gDNA calibrants. pDNA calibrants exhibited a lower bias in terms of closeness to the expected value of unknown samples than their genomic counterparts. The assay was specific and sensitive, where the limit of detection (LOD) and limit of quantification (LOQ) were five copies and ten copies per reaction. The short-term stability study of the pDNA calibrants indicated its stability for 60 days at − 20 °C and 30 days at 4 °C. The efficient results indicated a plasmid calibrator as a potential tool for absolute quantification of the pvalb gene and an alternative to conventional gDNA standards.
Czech name
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Czech description
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Classification
Type
J<sub>imp</sub> - Article in a specialist periodical, which is included in the Web of Science database
CEP classification
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OECD FORD branch
10608 - Biochemistry and molecular biology
Result continuities
Project
<a href="/en/project/QK1910231" target="_blank" >QK1910231: New approaches for the proof of fish meat adulteration using genomic DNA</a><br>
Continuities
P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)<br>I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace
Others
Publication year
2023
Confidentiality
S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů
Data specific for result type
Name of the periodical
European Food Research and Technology
ISSN
1438-2377
e-ISSN
1438-2385
Volume of the periodical
249
Issue of the periodical within the volume
12
Country of publishing house
DE - GERMANY
Number of pages
10
Pages from-to
3165–3174
UT code for WoS article
001089263300003
EID of the result in the Scopus database
2-s2.0-85168858155