An optimized TaqMan real-time PCR method for authentication of ASINI CORII COLLA (donkey-hide gelatin).
Abstract: In this study, probe/primers of high specificity and sensitivity were selected to analyze donkey-hide gelatin for donkey DNA and to look for horse, ox, and pig DNA as possible adulterants. The mitochondrial CO I genes in donkey, horse, and ox were selected as target sequences for design and synthesis of three pairs of specific probes and primers. In addition, eight pairs of probe/primers were obtained via literature search. Out of these eleven groups of probe/primers, those with the highest specificity and sensitivity were selected, which was fulfilled by the screening firstly with animal hide samples then the hide-glue samples. Other parameters that might affect detection specificity and efficiency-such as the amount of sampling and final concentration of primers-were also optimized. Replication tests were also conducted. The results showed that the selected probe/primers could accurately detect donkey DNA and horse, ox, and pig DNA in gelatin samples with good reproducibility. Analysis of four samples of on-market gelatin using this assay showed that two of the four samples indeed contained only donkey DNA, whereas the other two samples contained both donkey and horse DNA, indicating adulteration of these samples with horse hide. These results indicate that the TaqMan probe real-time PCR method can be used for identifying the purity of donkey DNA in gelatin samples, and can provide technical support for identifying adulterations in the gelatin market.
Copyright © 2019 Elsevier B.V. All rights reserved.
Publication Date: 2019-03-15 PubMed ID: 30928895DOI: 10.1016/j.jpba.2019.03.028Google Scholar: Lookup
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- Journal Article
Summary
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This research article focuses on enhancing a method using TaqMan real-time PCR for identifying if donkey-hide gelatin, also known as ASINI CORII COLLA, has been adulterated with DNA of horse, ox, or pig.
Research Objectives and Method
- The researchers aimed to authenticate donkey-hide gelatin (ASINI CORII COLLA) by identifying potential adulteration with horse, ox, and pig DNA.
- They selected probe/primers with high sensitivity and specificity to detect and analyze the donkey DNA present in the gelatin.
- For this purpose, the mitochondrial CO I genes in donkey, horse, and ox were chosen as target sequences, leading to the design and synthesis of specific probes and primers.
- Additionally, eight pairs of probe/primers from existing literature were taken into consideration.
Screening and Testing
- These eleven groups of probe/primers underwent screening, initially with animal hide samples followed by hide-glue samples.
- Screening aimed to identify probe/primers with the highest specificity and sensitivity for detecting DNA.
- The team also adjusted parameters such as the sampling amount and final concentration of primers to enhance detection efficiency and specificity.
- Replication tests were conducted to ensure the accuracy and validity of the chosen probe/primers.
Outcome and Market Relevance
- The chosen probe/primers demonstrated accurate detection of donkey DNA and horse, ox, and pig DNA in gelatin samples with high reproducibility.
- Using this method, four gelatin samples from the market were tested. Two were found to only contain donkey DNA, while the other two were adulterated with horse hide, that is, they contained both donkey and horse DNA.
- The results provide a clear indication that the optimized TaqMan real-time PCR method can verify the purity of donkey DNA in gelatin samples. This method can offer technical support for identifying adulterations in the gelatin market, ensuring the purity and authenticity of the products.
Cite This Article
APA
Zhang W, Cui S, Cheng XL, Wei F, Ma S.
(2019).
An optimized TaqMan real-time PCR method for authentication of ASINI CORII COLLA (donkey-hide gelatin).
J Pharm Biomed Anal, 170, 196-203.
https://doi.org/10.1016/j.jpba.2019.03.028 Publication
Researcher Affiliations
- National Institutes for Food and Drug Control, NO. 2, Tiantan Xili, Dongcheng District, Beijing, 100050, PR China. Electronic address: zhangwenjuan@nifdc.org.cn.
- National Institutes for Food and Drug Control, NO. 2, Tiantan Xili, Dongcheng District, Beijing, 100050, PR China.
- National Institutes for Food and Drug Control, NO. 2, Tiantan Xili, Dongcheng District, Beijing, 100050, PR China.
- National Institutes for Food and Drug Control, NO. 2, Tiantan Xili, Dongcheng District, Beijing, 100050, PR China. Electronic address: weifeng@nifdc.org.cn.
- National Institutes for Food and Drug Control, NO. 2, Tiantan Xili, Dongcheng District, Beijing, 100050, PR China. Electronic address: masc@nifdc.org.cn.
MeSH Terms
- Animals
- DNA / genetics
- DNA Primers / genetics
- Equidae / genetics
- Gelatin / genetics
- Horses
- Real-Time Polymerase Chain Reaction / methods
- Reproducibility of Results
- Sensitivity and Specificity
- Swine
Citations
This article has been cited 5 times.- Mahamad P, Dahlan W, Kahong S, So-Audon S, Munaowaroh W, Nopponpunth V. Duplex droplet digital PCR (ddPCR) for simultaneous quantification of bovine and porcine gelatin in capsules.. Food Sci Biotechnol 2023 May;32(6):803-811.
- Xia Z, Che X, Ye L, Zhao N, Guo D, Peng Y, Lin Y, Liu X. A Synergetic Strategy for Brand Characterization of Colla Corii Asini (Ejiao) by LIBS and NIR Combined with Partial Least Squares Discriminant Analysis.. Molecules 2023 Feb 13;28(4).
- Chen S, Li Z, Zhang S, Zhou Y, Xiao X, Cui P, Xu B, Zhao Q, Kong S, Dai Y. Emerging biotechnology applications in natural product and synthetic pharmaceutical analyses.. Acta Pharm Sin B 2022 Nov;12(11):4075-4097.
- Zhang J, Wu M, Ma Z, Zhang Y, Cao H. Species-specific identification of donkey-hide gelatin and its adulterants using marker peptides.. PLoS One 2022;17(8):e0273021.
- Bennett R, Pfuderer S. The Potential for New Donkey Farming Systems to Supply the Growing Demand for Hides.. Animals (Basel) 2020 Apr 20;10(4).
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