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1.
Wen‐Tao Xu Kun‐Lun Huang Ying Wang Hong‐Xing Zhang Yun‐Bo Luo 《Journal of the science of food and agriculture》2006,86(7):1103-1109
Biotechnology has permitted the modification of agricultural materials in a very precise way to improve productivity and yields. Polymerase chain reaction (PCR)‐based methods have been the first choice of most analytical laboratories for routine use in the detection of genetically modified organisms (GMO) and their derived products. These methods rely on the amplification of transgenic sequences and quantification of the transgenic DNA by comparison with an amplified reference gene. This paper describes the specific primers and probe for the cotton stearoyl‐ACP desaturase (sad1) gene, and PCR cycling conditions suitable for the use of this sequence, which acts as an endogenous reference gene in both qualitative and quantitative PCR assays. The two methods were tested with 18 cotton varieties and identical amplification products were obtained with all of them. No amplification products were detected when DNA samples from other species, including soybean, rapeseed, tobacco, maize, tomato, potato, cucumber, pea, red pepper, sunflower, sesame, rice, peach, banana, apple, pumpkin, barley and carrot, were used as templates, which demonstrates that this system is specific for cotton. In real‐time quantitative PCR analysis, the detection limit was as low as 6 pg of DNA, which indicates that this method is suitable for application to processed food samples that contain very low copies of target DNA. Southern blot analysis confirmed that the sad1 gene was a single copy in the tested cotton varieties. Copyright © 2006 Society of Chemical Industry 相似文献
2.
A real-time polymerase chain reaction (PCR)-based method for the detection of the walnut (Juglans regia) component in food is described here. The method consists of DNA isolation by chaotropic solid phase extraction and the subsequent PCR with walnut-specific primers and a TaqMan fluorescent probe. The primers and the probe are targeted to the jug r2, a major allergen gene of walnut. The method was positive for 8 varieties of walnut and negative for all other tested plant materials used in food industry, including pecan nuts. The intrinsic detection limit of the method was 0.24 ng walnut DNA. Using a series of model pastry samples with defined walnut contents, a practical detection limit of 0.01% walnut content was estimated. Practical applicability of the PCR method was tested by the analysis of 13 food samples (bakery and confectionery products), out of which two cakes were found to contain walnuts although they were not adequately labelled. The presented PCR method is useful for sensitive and selective detection of walnuts in food samples and can be performed in one working day. 相似文献
3.
A real-time PCR-based method for the detection of the pecan (Carya illinoiensis) component in food is described. The method consists of DNA isolation by chaotropic solid phase extraction and the subsequent
PCR with pecan-specific primers and a TaqMan fluorescent probe. The primers and the probe are targeted to the putative gene
for allergenic vicilin-like seed storage protein of pecan. The method was positive for 10 pecan varieties and negative for
all other tested plant materials used in food industry, including walnut. The intrinsic detection limit of the method was
1 pg pecan DNA which corresponds to 1.2 haploid genome copies. Using a series of model pastry samples with defined pecan contents,
a practical detection limit of 0.01% (w/w) pecan was estimated. Practical applicability of the PCR method was tested by the
analysis of 13 food samples; no discrepancies between the declared and detected pecan contents were found. The presented PCR
method is useful for sensitive and selective detection of pecans in food samples and can be performed in one working day. 相似文献
4.
A novel real-time polymerase chain reaction (PCR) method for the detection of hazelnuts in food 总被引:1,自引:0,他引:1
A real-time PCR (polymerase chain reaction)-based method for the detection of hazelnuts (nuts of Corylus avellana or C. maxima) in confectionery and bakery products is described. The method consists of DNA isolation by chaotropic solid phase extraction
and the subsequent PCR with hazelnut-specific primers and a TaqMan fluorescent probe. The primers and the probe are targeted
to the hsp1 gene encoding for a low molecular weight heat-shock protein. The method was positive for five hazelnut varieties approved
in Slovakia and negative for all other tested plant materials used in food industry including peanuts, walnuts, almonds, pistachio
nuts, cashews and chestnuts. The intrinsic detection limit of the method was 13 pg hazelnut DNA, which corresponds to approximately
27 genome equivalents (1C). Using a series of model pastry samples with defined hazelnut contents, a practical detection limit
of 0.01% (w/w) hazelnut was determined. Practical applicability of the PCR method was tested by the analysis of 20 food samples
(confectionery and bakery products) along with ELISA. For all of the food samples, identical results were obtained by both
methods, which conformed to the labelling. The presented PCR method is useful for sensitive and selective detection of hazelnuts
in food samples and can be performed in one working day. 相似文献
5.
A real-time PCR-based method for the detection of macadamia nuts (fruits of Macadamia integrifolia or M. tetraphylla or their hybrids) in food products is described. The method consists of DNA isolation by chaotropic solid phase extraction
and subsequent PCR with macadamia-specific primers and a TaqMan fluorescent probe. The primers and the probe were targeted
to the gene encoding for vicilin precursor. The method was positive for M. integrifolia and M. tetraphylla and negative for 16 other plant species used in food industry, including peanuts, walnuts, hazelnuts, almonds, pistachio
nuts, cashew nuts, Brazil nuts, and chestnuts. The DNA-based detection limit of the method was 1.45 pg. Using a series of
model samples with defined macadamia nut contents, a practical detection limit of 0.02% (w/w) macadamia nuts was determined.
Practical applicability of the PCR method was tested by the analysis of 14 confectionery samples. For all of the samples,
results conforming to the labeling were obtained. The presented PCR method is useful for relatively fast, highly selective,
and moderately sensitive detection of macadamia nuts in food samples. 相似文献
6.
In order to provide an appropriate method for the detection of pistachio (Pistacia vera) in food products, a novel real-time PCR was developed. The pistachio-specific primers and the TaqMan fluorogenic probe were
designed to target the internal transcribed spacer between 18S ribosomal RNA and 5.8S ribosomal RNA genes. Using dilutions
of the pistachio DNA, the intrinsic detection limit of the method was determined to be 0.012 pg. At specificity testing, the
method was positive for 11 pistachio varieties and negative for 26 plant and animal species used in food industry. A detection
limit of 0.0004% (w/w) was determined for pistachio nuts in model pastry. Practical applicability of the elaborated method was tested by the analysis
of 44 food samples, out of which 7 food products were identified as containing undeclared pistachio. The developed real-time
PCR may be utilized for sensitive and selective detection of pistachio in food products. 相似文献
7.
Rapid and Sensitive Detection of Staphylococcus aureus in Food Using Selective Enrichment and Real-Time PCR Targeting a New Gene Marker 总被引:1,自引:0,他引:1
Tereza Trnčíková Vendula Hrušková Katarína Oravcová Domenico Pangallo Eva Kaclíková 《Food Analytical Methods》2009,2(4):241-250
Staphylococcus aureus is a bacterial pathogen considered a principal etiological agent of food poisoning. The aim of this study was to develop
and evaluate a rapid and sensitive method for the detection of S. aureus in food by using selective enrichment and a new species-specific real-time polymerase chain reaction (PCR). Specific primers
and a TaqMan probe targeted to specific S. aureus gene encoding for acriflavine resistance protein were designed. The real-time PCR was highly specific for S. aureus with 100% inclusivity and 100% exclusivity determined using 83 S. aureus strains and 64 non-S.-aureus strains. PCR detection limit of 6.8 × 101 and 3.4 × 101 CFU ml−1 were obtained with 100% and 70% detection probability, respectively. The single selective enrichment based on the study of
different enrichment conditions was selected and a lysis by boiling was used to obtain bacterial DNA. Out of 112 food samples
analyzed, 61 were positive by the PCR-based method and 53 by the standard method. Out of ten food matrices artificially contaminated
at a level of 10° CFU g−1, ten and six were positive by the respective methods. Moreover, 10° CFU 10 g−1 was detected in all ten artificially contaminated samples after a large-scale enrichment using PCR-based detection, in contrast
to seven false negative by standard detection. The developed method facilitated the detection of S. aureus on the next day after the sample reception. This method can be used for S. aureus detection as a faster, highly specific, and more sensitive alternative to microbiological method with the potential for providing
of improved food-processing hygiene control. 相似文献
8.
Mohd Hazim Mohd Yusop Shuhaimi Mustafa Yaakob B. Che Man Abdul Rahman Omar Nur Fadhilah Khairil Mokhtar 《Food Analytical Methods》2012,5(3):422-429
Pork identification in raw meat using real-time polymerase chain reaction (PCR) was developed. Total DNA from meat samples
were successfully extracted and found to be of high quality and produced clear PCR products. Porcine-specific molecular beacon
probe and primers that amplifies 119 bp of the cytochrome b gene fragment of swine (Sus scrofa domestica) was used. Analysis of data showed that the C
q (quantification cycle) from 10 ng/μl porcine DNA is (18.70 ± 0.12 to 19.08 ± 0.06). Meanwhile, the other samples exhibited
negative result, which confirmed the specificity of the primers. The method also showed that the limit of detection of pork
was 0.0001 ng. Based on the regression analysis of the standard curve, the 96% efficiency of real-time PCR was achieved with
high correlation coefficient (r
2 = 0.9989). Sensitivity of the assay in discriminating pork as low as 0.1% (w/w) pork in pork–beef mixtures was also obtained. Reproducibility of the assay was successfully validated by applying sample
and experimental replicates in every assay being conducted. Thus, this methodology could serve as a fast and sensitive method
for detection of pork for meat species verification. 相似文献
9.
Michaela Petrášová Bohuslava Tremlová Zdeňka Javůrková 《Food additives & contaminants. Part A, Chemistry, analysis, control, exposure & risk assessment》2016,33(8):1283-1289
In this study we developed an immunofluorescence method to detect pea protein in meat products. Pea protein has a high nutritional value but in sensitive individuals it may be responsible for causing allergic reactions. We produced model meat products with various additions of pea protein and flour; the detection limit (LOD) of the method for pea flour was 0.5% addition, and for pea protein it was 0.001% addition. The repeatabilities and reproducibilities for samples both positive and negative for pea protein were all 100%. In a blind test with model products and commercial samples, there was no statistically significant difference (p > 0.05) between the declared concentrations of pea protein and flour and the immunofluorescence method results. Sensitivity was 1.06 and specificity was 1.00. These results show that the immunofluorescence method is suitable for the detection of pea protein in meat products. 相似文献
10.
M. Eaqub Ali U. Hashim Th. Sabar Dhahi S. Mustafa Yaakob Bin Che Man Md. Abdul Latif 《Food Analytical Methods》2012,5(4):784-794
A TaqMan probe real-time polymerase chain reaction assay was developed for the determination of pork adulteration in commercial
burgers. The assay combined porcine-specific primers and TaqMan probe for the selective amplification and detection of a 109-bp
fragment of swine cytochrome b (cytb) gene. Specificity test with 10 ng DNA of 11 different meat-providing animal and fish
species yielded a quantification cycle (Cq) of 15.5 ± 0.20 for the pork and negative results for the others in a 40-cycle
reaction with a change of analysts and sources. Analysis of beef burger formulations with spiked pork showed the assay can
determine 100–0.01% contaminated pork with a PCR efficiency (E) of 93.8% and a correlation coefficient (R
2) of 0.991. A plot of actual value against real-time PCR-predicted value also yielded a good linear regression, R
2 0.998, and small root mean square error of calibration, RMSEC 0.42. A strong correlation was found between the partial least square (PLS)-predicted values and real-time PCR-determined
values. The accuracy of the method was ≥90% in all determinations of the standard set. Residual analysis also revealed a high
precision in all determinations. Finally, a random analysis of 10 ng DNA of commercial burgers from pork, beef, chicken, mutton,
and chevon yielded a Cq of 15.56 ± 0.22 to 16.24 ± 0.35 from pork burgers, and negative results from the others, showing the
suitability of the assay to determine pork in commercial burgers with a high accuracy and precision. 相似文献
11.
12.
BACKGROUND: A one‐step polymerase chain reaction (PCR) method for the simultaneous detection of the major allergens of pecan and Brazil nuts was developed. Primer pairs for the amplification of partial sequences of genes encoding the allergens were designed and tested for their specificity on a range of food components. RESULTS: The targeted amplicon size was 173 bp of Ber e 1 gene of Brazil nuts and 72 bp of vicilin‐like seed storage protein gene in pecan nuts. The primer pair detecting the noncoding region of the chloroplast DNA was used as the internal control of amplification. The intrinsic detection limit of the PCR method was 100 pg mL?1 pecan or Brazil nuts DNA. The practical detection limit was 0.1% w/w (1 g kg?1). The method was applied for the investigation of 63 samples with the declaration of pecans, Brazil nuts, other different nut species or nuts generally. In 15 food samples pecans and Brazil nuts allergens were identified in the conformity with the food declaration. CONCLUSION: The presented multiplex PCR method is specific enough and can be used as a fast approach for the detection of major allergens of pecan or Brazil nuts in food. Copyright © 2011 Society of Chemical Industry 相似文献
13.
Claudia Unterberger Florian Luber Anja Demmel Karola Grünwald Ingrid Huber Karl-Heinz Engel Ulrich Busch 《European Food Research and Technology》2014,239(4):559-566
People suffering from food allergy rely on correct food labelling as the ingestion of minimal amounts of the respective allergen can trigger severe allergenic reactions. Probes for the detection of DNA from allergenic fish, shellfish and cephalopod species in food using multiplex ligation-dependent probe amplification were developed. The specificity and the sensitivity of the detection system were investigated. The limit of detection was 20 mg kg?1 for scallop, fish and bivalve species and 100 mg kg?1 for cephalopod, gastropod and crustacean species using self-prepared sushi spiked with the analytes in different concentration levels. The analysis of 10 commercial food samples demonstrates the applicability of the developed method and its suitability for food quality control. Therefore, the method can be used to monitor the compliance with labelling rules regarding food allergens. 相似文献
14.
A method based upon polymerase chain reaction (PCR) for the detection of celery (Apium graveolens) in food was developed. The method involves DNA isolation by chaotropic or non-chaotropic solid-phase extraction and PCR with primers oriented to the sequence of the nuclear gene encoding mannitol dehydrogenase. The PCR method was shown to be specific for celery, producing a 279 bp fragment with four celery varieties and negative results with other species commonly present together with celery in food products (16 samples). The detection limit of PCR was 490–1530 pg DNA, which corresponds to 102 genome copies. When evaluated with model samples of celery in meat pâtés, a detection limit of 0.1% (w/w) was determined. When used to analyse food products from the market (dried vegetable seasonings, dehydrated bouillons), all four products declared to contain celery were correctly identified as positive and all three products in which celery was not declared were identified as negative. 相似文献
15.
Detection of eight GMO maize events by qualitative, multiplex PCR and fluorescence capillary gel electrophoresis 总被引:1,自引:1,他引:0
Specific legislation in the EU and several other countries requires that foods containing genetically modified organisms (GMOs)
should be approved and labelled. This has necessitated the development of methods for detection of such materials. For screening
purposes these methods should preferably enable detection of several different GMOs. Here we present a simple, robust, qualitative,
nineplex PCR method for event-specific detection of maize T25, GA21, TC1507, MON863, MON810, NK603, construct specific detection
of BT176, BT11 and detection of the endogenous hmga maize reference gene. PCR is carried out with primers labelled with fluorescent groups and the amplicons are detected using
fluorescence capillary electrophoresis. Using mixtures of DNA from different certified reference materials, the detection
limit was determined to approximately 0.1% for each GMO. Good agreement was observed in 85 of 88 determinations when eleven
food and feed samples were analysed using the multiplex PCR assay and compared to results from quantitative real-time 5′-nuclease
PCR. Discrepancies were only observed for one GMO at or close to the detection limit. The presented method is therefore suitable
for screening purposes for food and feed containing the most common maize GMOs. 相似文献
16.
St. Maryam Sismindari Tri Joko Raharjo Sudjadi Abdul Rohman 《International Journal of Food Properties》2016,19(1):187-195
The identifications of species in meat products have created interests since these foods became the target of forgery and fraud in the market. The presence of pork in food products is not allowed for the Muslim community. Hence, an analysis is necessary to detect the presence of pork in processed meat products, such as in dendeng (dried meat) product. Real time polymerase chain reaction using mitochondrial displacement loop686 and cytochrome b (cytb) gene primers was used to identify specific pork DNA among other four types of DNA species; namely beef, chicken, goat, and horse. This method was also used to identify pork DNA in the laboratory processed pork-beef dendeng as well as commercial dendeng from market. The results showed that real time polymerase chain reaction using displacement loop686 and cytb gene primers were able specifically to distinguish between pork DNA and the other species. The lowest concentration of 0.5% of pork DNA in a mixture of pork-beef processed products of dendeng was able to be detected by both primers with the product amplification of 114 and 134 bp (base pair) for the displacement loop686 and 149 bp for cytb gene, respectively. High sensitivity was also obtained when both primers were applied with the lowest detection limit of 5 pg/µL pork DNA. The results of the six commercial dendeng amplification using both primers showed no amplified products present, meaning that these products do not contain porcine DNA. 相似文献
17.
Detection of Salmonella spp., Yersinia enterocolitica,Listeria monocytogenes and Campylobacter spp. by real‐time multiplex PCR using amplicon DNA melting analysis and probe‐based assay 下载免费PDF全文
Jūratė Skerniškytė Julija Armalytė Raimonda Kvietkauskaitė Vaida Šeputienė Justas Povilonis Edita Sužiedėlienė 《International Journal of Food Science & Technology》2016,51(2):519-529
Syto9 and probe‐based multiplex real‐time PCR assays for simultaneous detection of a group of foodborne pathogens (named SYLC group), targeting Salmonella spp. (invA gene), Yersinia enterocolitica (ystA gene), Listeria monocytogenes (hly gene) and Campylobacter spp. (rrna gene), have been developed. The Syto9 assay generates amplicon DNA melting curve with four peaks of 86.5 ± 0.5, 84 ± 0.5, 81.5 ± 0.5 and 90.5 ± 0.5 °C corresponding Salmonella spp., Y. enterocolitica, L. monocytogenes and Campylobacter spp. targets, respectively. The sensitivities of the Syto9 and TaqMan assays in artificially inoculated chicken wing rinses were in a range of 3.2 × 102 to 3.1 × 104 and 9.8 × 102 to 1.9 × 104 colony‐forming units per millilitre, respectively, depending on the pathogen. All tested target strains (n = 100) were correctly detected by the both assays, whereas nontarget strains (n = 100) demonstrated no cross‐reactivity representing 100% specificity. The assays are suitable for application in qualitative and quantitative detection of SYLC group pathogens in food matrices. 相似文献
18.
《Journal of dairy science》2022,105(12):9450-9462
Foodborne pathogens detection is important to ensure food safety and human health. In this study, we designed a comet structure to rapidly and sensitively detect foodborne Listeria monocytogenes. This method combined isothermal sequence exchange amplification (SEA) and surface-enhanced Raman spectroscopy. Listeria monocytogenes DNA could be rapidly amplified at a constant temperature via SEA with a pair of modified primers, which rendered the precise thermal control instrumentation unnecessary. Efficient SEA amplification generated a large number of DNA duplexes that could be easily captured by streptavidin-modified magnetic bead and AuMB@Ag-isothiocyanate fluorescein antibody (anti-FITC). AuMB@Ag-anti-FITC was used as a signal probe, which generated a significant excitation signal at 1,616 cm?1 for quantitative detection and analysis. The results displayed sensitive detection of L. monocytogenes in cheese from 2.0 × 101 cfu/mL to 2.0 × 106 cfu/mL within 1.0 h with a detection limit of 7.8 cfu/mL. Furthermore, this comet structure displayed the desirable specificity as its specific primers and amplified DNA ends were attached to streptavidin-modified magnetic beads and AuMB@Ag-anti-FITC, respectively. We expected that the method devised would provide a promising new approach to screening for L. monocytogenes and guarantee the microbiological safety of dairy products. 相似文献
19.
Asing Sharifah Bee Abd Hamid Motalib Hossain Mohammad Nasir Uddin Ahamad S. M. Azad Hossain 《Food additives & contaminants. Part A, Chemistry, analysis, control, exposure & risk assessment》2016,33(11):1643-1659
The Malayan box turtle (Cuora amboinensis) (MBT) is a vulnerable and protected species widely used in exotic foods and traditional medicines. Currently available polymerase chain reaction (PCR) assays to identify MBT lack automation and involve long targets which break down in processed or denatured tissue. This SYBR Green duplex real-time PCR assay has addressed this research gap for the first time through the combination of 120- and 141-bp targets from MBT and eukaryotes for the quantitative detection of MBT DNA in food chain and herbal medicinal preparations. This authentication ensures better security through automation, internal control and short targets that were stable under the processing treatments of foods and medicines. A melting curve clearly demonstrated two peaks at 74.63 ± 0.22 and 78.40 ± 0.31°C for the MBT and eukaryotic products, respectively, under pure, admixed and commercial food matrices. Analysis of 125 reference samples reflected a target recovery of 93.25–153.00%, PCR efficiency of 99–100% and limit of detection of 0.001% under various matrices. The quantification limits were 0.00001, 0.00170 ± 0.00012, 0.00228 ± 0.00029, 0.00198 ± 0.00036 and 0.00191 ± 0.00043 ng DNA for the pure meat, binary mixtures, meatball, burger and frankfurter products, respectively. The assay was used to screen 100 commercial samples of traditional Chinese herbal jelly powder from eight different brands; 22% of them were found to be MBT-positive (5.37 ± 0.50–7.00 ± 0.34% w/w), which was reflected through the Ct values (26.37 ± 0.32–28.90 ± 0.42) and melting curves (74.63–78.65 ± 0.22°C) of the amplified MBT target (120 bp), confirming the speculation that MBT materials are widely used in Chinese herbal desserts, exotic dishes consumed with the hope of prolonging life and youth. 相似文献
20.
Maninder Meenu Anupma Sharma Paramita Guha 《International Journal of Food Properties》2016,19(10):2223-2237
A fast method was developed for simultaneous detection and quantification of 12 phenolic compounds in mung bean, using reversed phase high-performance liquid chromatography. The method was optimized for mobile phase combination, elution gradient, detection wavelength, and solvent extraction. All the phenolic compounds (gallic acid, neochlorogenic acid, catechin, chlorogenic acid, caffeic acid, p-coumaric acid, t-ferulic acid, vitexin, isovitexin, myricetin, quercetin, and kaempferol) were eluted for 18 min and recovered within a limit as per International Council for Harmonization guidelines. The method showed good linearity with correlation coefficient of 0.998. The limit of detection and quantification of all the compounds ranges from 0.27 ± 0.01 to 3.65 ± 0.3µg/mL and 0.91 ± 0.1 to 12.17 ± 0.9µg/mL, respectively. Vitexin (28.10 ± 0.20 to 29.60 ± 0.6 mg/100 g raw material) and isovitexin (34.09 ± 0.14 to 36.83 ± 0.82 mg/100 g raw material) were the major phenolic compounds along with other phenolic compounds found in mung beans. 相似文献