Detection of genetically modified rice: collaborative validation study of a construct-specific real-time PCR method for detection of transgenic Bt rice

A collaborative trial study has been conducted for validation of an extraction method and a subsequent real-time PCR for detection of a transgenic Bt rice line (‘Bt63’) in rice products originating from China. A total of 17 laboratories participated in the study and each laboratory received 16 coded samples comprising of rice grain flours, rice noodle flours and plasmid DNAs. Of the accepted results all Bt63-positive rice grain samples (0.1 or 0.05% w/w) and all rice noodle samples prepared from marketed rice products were detected correctly. The result demonstrates that ‘Bt63’ rice is detectable even at low relative mass concentrations of 0.05%. The absolute LOD determined with plasmid DNA samples showed to be at least five copies of the ‘Bt63’ target sequence. The data provided in this study show that the method is fit-for-purpose to inspect Chinese rice products for the presence of EU-unauthorised rice lines carrying the ‘Bt63’ construct.     

Identification and quantification of anthocyanins from the grains of black rice (Oryza sativa L.) varieties

The purpose of this research was conducted to determine anthocyanin content from the grains of 10 Korean black rice varieties. Moreover, the primary constituents including protein and oil were evaluated. Anthocyanins, cyanidin-3-O-glucoside (1) and peonidin-3-O-glucoside (2), were isolated and elucidated using reversephase C₁₈ chromatography, nuclear magnetic resonance (NMR) spectroscopy, and high performance liquid chromatography (HPLC) with diode array detection and electrospray ionization/mass spectrometry (DAD-ESI/MS). Anthocyanin showed significant differences and cyanidin-3-O-glucoside (1) (52.1±6.3-1,601.0±8.5 μg/g) exhibited a markedly higher content than peonidin-3-O-glucoside (2) (ND-82.6±1.2 μg/g). Among varieties, ‘Heugjinjubyeo’ showed the highest anthocyanin (1: 1,601.0±8.5, 2: 82.6±1.2 μg/g), whereas ‘Heughyang’ was the lowest (1: 52.1±6.3 μg/g, 2: ND). Protein and oil exhibited the minor differences and their contents ranged from 10.7±0.0 to 14.1±0.1% and from 2.1±0.0 to 2.9±0.0%. Overall results suggest that anthocyanin can be a key factor in functional property of black rice and ‘Heugjinjubyeo’ may be very important source concerning nutritional value.     

转基因水稻Bt63的外源基因相对含量分析

为分析转基因Bt63稻米外源基因含量,利用新型、灵敏和高通量的实时荧光定量PCR技术,以RBE-4作为转基因水稻的内源参照基因,通过梯度稀释法,分别获得了Bt 63和RBE-4基因的Ct值与起始模板相关性的标准曲线,相关系数分别为0.996 6和0.995 4。通过转基因水稻外源基因(Bt63)和内标基因(RBE-4)起始模板数的比较,测定了外源基因在转基因水稻中的相对含量,这项研究将为建立转基因水稻抗虫能力评价体系和转基因稻谷的流通和监管提供技术支撑。     

Development of an event-specific Real-time PCR detection method for the transgenic Bt rice line KMD1

The present study describes an event-specific quantitative Real-time PCR detection method for the transgenic Bt rice line Kemingdao 1 (KMD1). This rice line which is not approved in any country so far is likely to be approved in China in the near future. The developed primers amplify a DNA sequence spanning the integration site of the genetic construct in KMD1. DNA sequence information of this unknown site necessary for primer design was achieved using SiteFinding-PCR technique. The specificity of the detection method was shown by testing a number of different transgenic and conventional plant varieties (e.g. rice LL 601, LL 62, Bt 63). As alternative to genomic DNA, we developed double target hybrid amplicons as synthetic calibration standards in Real-time PCR. These amplicons contained both one copy of the KMD1 event-specific sequence and one copy of a sequence of the rice reference gene gos9. The limit of quantification (LOQ) of the method was tested to be 0.05%. R. Babekova and T. Funk contributed equally to this paper.     

Qualitative detection and quantification of a <Emphasis Type="Italic">Cry1A(b)</Emphasis> transgene present in rice cv. Zhejing 22

The rice cultivar Zhejing 22 has been transformed to carry the Bacillus thuringiensis Cry1A(b) gene in 2007. Although the cultivation of this line (referred to as Bt-ZJ22) is yet to be authorized, it is likely to be approved in China in the near future. The transgene-host 3’ integration junction was isolated using thermal asymmetric interlaced PCR, and its sequence was used to design PCR primers and an appropriate TaqMan probe for a real-time PCR quantitative assay of the transgene. The limit of detection for a standard PCR assay was estimated at ten gene copies, while that for the real-time PCR assay was 5–10 copies. The latter assay was tested on samples of rice DNA with either 5, 3, 1, or 0.5% Bt-ZJ22; this produced estimates for the relative content of Bt-ZJ22 of, respectively, 5.3, 3.2, 0.97, and 0.48%. The precision of these estimates demonstrated that this real-time PCR assay should be suitable for the detection and monitoring of this transgenic event.     

Combinatory SYBR® Green Real-Time PCR Screening Approach for Tracing Materials Derived from Genetically Modified Rice

Two real-time PCR approaches for the detection of genetically modified (GM) rice were tested: (1) a combination of SYBR® Green real-time PCR methods detecting the 35S promoter (P-35S) of Cauliflower Mosaic Virus, the nopaline synthase terminator (T-nos) of Agrobacterium tumefaciens and the Bacillus thuringiensis (Bt) CryIAb/Ac toxins and (2) a P-35S/T-nos duplex TaqMan® real-time PCR system. Both systems correctly recognized their respective targets in control samples of Bt63, Kefeng6 and KMD1 insect-resistant and LLRice62 and LLRice601 herbicide-resistant rice. Due to its lesser specificity but broader genetically modified organism (GMO) coverage capacity, the SYBR® Green real-time PCR approach was further tested in more detail. Melting curve, capillary and agarose gel electrophoresis analyses indicated that the P-35S, T-nos and CryIAb/Ac targets in the GM rice events are similar to the corresponding targets present in many known GMOs. High-resolution melting analysis showed that the CryIAb/Ac targets of the GM rice events Bt63 and Kefeng6 matched best the corresponding Bt11 CryIAb sequence. Digital PCR analysis on genomic DNA from the GM rice Bt63 and Kefeng6 events indicated that both GMO contained multiple inserts. Sensitivity tests showed that all SYBR® Green real-time PCR methods could detect their targets at less than an estimated five copies per reaction. Finally, it was shown that these SYBR® Green real-time PCR methods could detect low levels of their targets in rice consignments originating from China. Together, these results demonstrated that a ‘P-35S and T-nos and CryIAb/Ac’ combinatory SYBR® Green real-time PCR screening is highly suited to trace the respective targets including the possible presence of Bt63, Kefeng6 and KMD1 GM rice materials in food products.     

Properties and qualities of rice flours and gluten-free cupcakes made with higher-yield rice varieties in Korea

Korean rice varieties, ‘Druryechanbyeo’ and ‘Boramchanbyeo’, were developed to get higher yields and to be used in rice products. The rice grains were dried and milled into rice flours after first going through the soaking process. The properties and qualities of cupcakes made with dry-milled rice flours were compared with cupcakes made with commercial dry-milled rice flours (CDRF). The newly developed rice flours (NDRF) had higher apparent amylose content, water binding capacity, swelling power, and peak viscosities, but had lower damaged-starch content, gelatinization temperature, and final and setback viscosities than CDRF. The specific gravity of batter, and hardness and springiness of cupcakes were lower in NDRF than in CDRF. The cake textures from ‘Boramchan’ NDRF were more preferable than those from ‘Druyechan’ NDRF. The specific volume and overall quality of cupcake were correlated positively with amylose content and water binding capacity, but negatively with damaged starch of rice flours.     

Development and validation of microscopical diagnostics for ‘Tulsi’ (<Emphasis Type="Italic">Ocimum tenuiflorum</Emphasis> L.) in ayurvedic preparations

During recent years, ayurvedic plants have entered the European market as a novel food trend. This confronts food analytics with the task to assess the composition of exotic and often unknown herbal preparations in teas or spices. Using the trend plant ‘Tulsi’ (Holy Basil, Ocimum tenuiflorum L.) as model, we developed microscopical diagnostics on markers that can be reliably assessed in dried or even fragmented specimens as typically occurring in commercial ayurvedic preparations, where DNA extraction is difficult. First, a reference for ‘Tulsi’ was defined based on the plastidic internal transcribed spacer (ITS) as marker. Second, this reference was morphologically delineated from other Ocimum accessions potentially used as surrogates for ‘Tulsi’ (such as O. basilicum L.) leading to a microscopical assay based on the density of glandular scales and glandular hairs, the epidermis with trichomes and the cells of the palisade parenchyma. Third, this assay was statistically validated for its ability to discriminate surrogate species from true O. tenuiflorum. First applications of this assay on commercial ‘Tulsi’ products demonstrated a high frequency of surrogate additions.     

Development of an event-specific detection method for genetically modified rice Kefeng?6 by quantitative real-time PCR

The genetically modified (GM) rice Kefeng 6 has gained resistance against several rice pests by inserting the cpti and cry1Ac genes. As this transgenic line is not approved for import, processing and cultivation in the European Union (EU), sensitive and specific detection methods need to be available to monitor any illegal presence of Kefeng 6 in food products within the EU. The aim of this study was to develop and validate an event-specific detection method by means of quantitative real-time PCR (qPCR) for the detection of Kefeng 6 in foodstuff. A primer pair and hydrolysis probe were designed according to the right border junction sequence of the transgene. The qPCR assay was validated according to the ENGL/EURL-GMFF guidelines for GMO testing and is presented according to the MIQE guidelines. The in-house validation process resulted in a limit of detection of 5 DNA copies of the transgene with confidence intervals (95 %) between 0.07 and 0.52, a PCR efficiency of 105 % and a correlation coefficient (R ²) value of 0.9997. The specificity of the assay was tested by end-point PCR, gel electrophoresis and subsequent sequencing of the PCR products. By testing DNA of several GM and non-GM crops, cross reactivity of the assay was not observed. Further, 35 food products were analyzed for the presence of Kefeng 6 by means of the event-specific detection method. For 9 out of 35 samples, PCR products for Kefeng 6 DNA were observed.     

Integrated structure and event-specific real-time detection of transgenic <Emphasis Type="Italic">cry1Ac/SCK</Emphasis> rice Kefeng 6

Transgenic rice Kefeng 6 is a transformation event containing two insect-resistant genes, cry1Ac and SCK (modified CpTI gene) in China. In order to monitor the probable release of Kefeng 6 in the future and execute the labeling requirements, it is necessary to develop a rapid and reliable detection method. In this study, both the 5′ and 3′-junction sequences spanning the plant DNA and the integrated gene construct of the rice event Kefeng 6 were isolated by genome walking and long-distance PCR (LD-PCR), successively. Multiple copies of truncated SCK gene and cry1Ac gene were found to integrate into the host rice genome. The event-specific real-time detection method for Kefeng 6 event based on its 5′-junction sequence was established using one plasmid molecule pMD-KF6 containing both 5′-junction sequence and rice endogenous gene gos9 sequence as the reference material (RM) with an absolute limit of quantification (LOQa) around 10 template copies. Thereafter, three different transgenic amounts of w/w mixed samples (5, 1, and 0.5%, respectively) were quantified to assess the performance characteristics of the established real-time PCR method. The accuracy expressed as bias deviated from the 4.00–26.00%, the precision expressed as standard deviation (SD) and relative standard deviation (RSD) deviated from 0.03–0.19 and 3.42–4.76%, respectively. Based on the earlier results, we concluded that the qualitative and quantitative PCR assays were reliable and accurate for Kefeng 6 measurement, and the reference plasmid pMD-KF6 could be a good substitute for the reference material for Kefeng 6 quantification.     

Collaborative trial validation of a construct-specific real-time PCR method for detection of genetically modified linseed event ‘CDC Triffid’ FP967

A real-time PCR-based method for construct-specific detection of the genetically modified (GM) linseed event ’CDC Triffid’ FP967 originating from Canada has been validated in a collaborative trial. The construct-specific method amplifies a 105 bp long fragment of the transgenic insertion present in FP967 spanning the junction of the terminator region of the nopalin synthase gene from Agrobacterium tumefaciens (Tnos) to a sequence region coding for the dehydrofolate reductase gene (dfr) from a class I integron from Escherichia coli. This region is characteristic for the construct used to develop FP967. A total of 11 laboratories participated in the collaborative study. For PCR analysis, each laboratory received 14 DNA samples comprising 7 double-blind DNA samples. The samples consisted of two low GM-levels of FP967 DNA (10 or 50 copies per PCR), of DNA from two different GM-positive linseed products and of DNA from GM-negative linseed, potato and rapeseed materials, respectively. All but one of the FP967-positive DNA samples were detected correctly. No false-positive results were reported. The results demonstrate that the linseed event FP967 is detectable even at low copy number concentrations. The limit of detection (LOD) determined with plasmid DNA was shown to be at 5 copies of the Tnos–dfr sequence. The data provided show that the method can be applied successfully in different laboratories and is fit-for-purpose to test for the presence of the EU-unauthorised linseed event ‘CDC Triffid’ FP967.     

A papaya-specific gene, papain, used as an endogenous reference gene in qualitative and real-time quantitative PCR detection of transgenic papayas

Genetically modified (GM) papaya lines have been approved for commercialization and are widely cultivated in many countries. As a step towards the development of reliable qualitative and quantitative PCR methods for detecting GM papayas, one papaya (Fructus Caricae Carica papaya L.) species specific gene, papain, was selected and validated as suitable for using as an endogenous reference gene in transgenic papaya PCR detection. In this article, both qualitative and quantitative PCR assays for the papain gene were assayed with 11 different papaya varieties and identical amplification products were obtained with all of them. No amplified fragments could be detected when DNA samples from 18 kinds of other species were used as templates, which demonstrated that this system was specific for papaya. In real-time quantitative PCR analysis, the detection limit was as low as 10 pg of DNA. Southern blot analysis confirmed that the papain gene was two copies in the tested papaya varieties and no allelic variation was testified among these tested papaya varieties. In addition, the common used exogenous genes of the coat protein (CP) and the replicase (RP) were also assayed in qualitative and real-time quantitative PCR. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Wentao Xu and Weibin Bai contributed equally to this work.     

Toxicological evaluation of transgenic rice flour with a synthetic cry1Ab gene from Bacillus thuringiensis

Bacillus thuringiensis (Bt) transgenic (KMD1) and non‐transgenic (KMD1′s parental variety Xiushui 11) rice flours were assessed in a 90 day feeding test with rats. KMD1 contained a synthetic cry1Ab gene from Bt, and selection marker genes nptII and hpt linked in tandem. In the≤64 g kg^?1 body weight (BW) dosage range (Bt transgenic rice flour composed 64% of the ingredients of the diet), no adverse effects of Bt rice on rats were observed in terms of animal behaviour, weight gain and feed utilisation rate. Necropsy at the end of the experiment indicated that neither pathological lesions nor histopathological abnormalities were present in organs such as liver, kidneys, intestines and testes of rats in both test and control groups by macroscopic and microscopic pathology. In addition, no significant differences (P > 0.05) were observed in relative organ weights, haemograms and blood indices of rats between test and control groups. Several serum parameters of female rats were found to be significantly different between Bt and non‐Bt diets, but the values of these parameters were still within the normal ranges of values for rats of this age and sex. These results demonstrated that Bt rice flour at a dosage of 64 g kg^?1 BW, Bt toxin and NPTII and HPT proteins have no toxic effects on rats. © 2002 Society of Chemical Industry     

Detection of Pathogenic Yersinia enterocolitica in Meat using Real-Time PCR

Yersinia enterocolitica is an important zoonosis, which can cause disease in humans and animals. The culture methods available for detection of Y. enterocolitica in food samples are time-consuming and seldom successful. Using DNA-based methods, like PCR, this pathogen can be detected more rapidly and with greater sensitivity. The aim of this study was to establish a rapid and accurate real-time PCR method to detect pathogenic Y. enterocolitica in pork. The chromosomal ail−gene, which is only present in pathogenic Y. enterocolitica strains, was used as DNA target for the 5' nuclease PCR assay. The probe was labelled at the 5' end with the fluorescent reporter dye (FAM) and at the 3' end with the quencher dye (TAMRA). A 2-step protocol with 45 cycles was used in a multicolour real-time PCR detection system. A Ct value over 40 indicated a negative result. The DNA extraction procedure for the natural samples was rapid and simply. This qualitative real-time PCR method was shown to be specific and sensitive. Detection rate of ail-positive Y. enterocolitica in 200 pig tonsils was 88 % and 35 % with PCR and culture methods, respectively. When 100 raw pork samples were studied, 7 were positive with PCR and all were culture negative. Received: January 30, 2007; Received in revised form: February 27, 2007; Accepted: February 28, 2007.     

Improved Sample Preparation for Real-Time PCR Detection of Listeria monocytogenes in Hot-Smoked Salmon using Filtering and Immunomagnetic Separation Techniques

This work evaluated the application of filtration and immunomagnetic separation (IMS) as sample pretreatments for use in combination with real-time polymerase chain reaction (PCR) to detect and quantify Listeria monocytogenes in hot-smoked salmon. Salmon was artificially inoculated with L. monocytogenes at levels ranging from 8 × 10⁰ to 8 × 10⁵ cfu/g of sample, and homogenates obtained from these samples were filtered to recover bacterial cells without a pre-enrichment step. High recovery of bacterial cells was achieved using standard coffee filters. IMS significantly reduced the co-extraction of PCR inhibitors present in the samples to increase the assay sensitivity with regression line parameters applicable for quantification. The limit of detection and quantification were equal to 2 × 10¹–4 × 10¹ and 2 × 10² cfu/g of sample, respectively. The entire detection procedure could be completed within 3.5 h. This study demonstrated that coupling filtration and IMS with real-time PCR has contributed to improve the sensitivity of L. monocytogenes detection from hot-smoked salmon.     

Inhibition of lipase from rice (Oryza sativa) by diethyl-p-nitrophenyl phosphate

The inhibition of rice bran lipase (RBL) by diethyl-p-nitrophenyl phosphate was studied with reference to kinetics, nature of inhibition and also elucidate the effect of the inhibitor on the structure—function of the enzyme. Enzyme activity measurements shows that the inhibitor is more effective at 0.050 mM concentration of diethyl-p-nitrophenyl phosphate and the activity is 50% at this level of inhibitor concentration. The affinity of substrate for the enzyme was observed by the increase in the velocity of the reaction with increase in the substrate concentrations and double reciprocal plot indicates that the inhibition followed a competitive in nature and inhibition constant K _i is found to be 0.016 mM at pH 7.0. The decrease in apparent thermal denaturation temperature to 4 °C compared to control indicates the destabilization of enzyme in the presence of inhibitor. Fluorescence spectral measurements suggests that pronounced quenching of fluorescence intensity of RBL occurs at higher concentrations of diethyl-p-nitrophenyl phosphate and ‘K _a’ value was found to be 2.4 × 10⁴ M⁻¹ with free energy change ΔG^o—26 kJ/mol at 30 °C suggesting strong binding between the enzyme and the inhibitor with microenvironmental changes occur at the active site or in the neighbourhood of active site. The far UV-CD data suggest that there is no significant changes in the conformation of the enzyme as a result of binding of diethyl-p-nitrophenyl phosphate. These results indicate that diethyl-p-nitrophenyl phosphate is a inhibitor of RBL and binds to the enzyme in brining about inhibition without any structural alterations.     

PCR-based quantification of genetically modified Bt maize: single-competitive versus dual-competitive approach

 Methods for the quantification of transgenic insect-resistant Bt maize by single- and dual-competitive PCR were developed. The analysis of mixtures of DNA solutions, as well as of maize flours containing defined amounts of Bt maize, demonstrated the usefulness of single-competitive PCR based on coamplification of the CDPK promoter/cryIA(b) gene region of Bt maize and an internal standard. Upon heat treatment of DNA solutions and maize flour, respectively, the recovery of the Bt proportion compared to the starting material determined by single-competitive PCR decreased significantly. This systematic error could be compensated for by using a dual-competitive approach based on PCR quantification of the transgenic target sequence of Bt maize and of the maize specific invertase gene (ivr1). Received: 24 January 2000 / Revised version: 25 February 2000     

Impact of minimal heat-processing on pectin methylesterase and peroxidase activity in freshly squeezed <Emphasis Type="Italic">Citrus</Emphasis> juices

The impact of minimal heat-processing of juices on the activities of endogenous pectin methylesterase (PE) and peroxidase (POD) was compared between Citrus species. Mono-cultivar juices were produced from three orange (Citrus sinensis (L.) Osbeck cvs. ‘Navelina’, ‘Salustiana’, and ‘Navelate’), two lemon (Citrus limon (L.) Burm. f. cvs. ‘Verna’ and ‘Primofiori’), and two Clementine mandarin varieties (Citrus reticulata Blanco cvs. ‘Marisol’ and ‘Clemenules’). Two mandarin hybrids (cvs. ‘Ortanique’ and ‘Clemenvilla’) were likewise used. The freshly squeezed juices were subjected to continuous treatments at six different temperatures (42–92 °C) with subsequent re-cooling on the pilot plant scale. In fresh Citrus juice, POD activities notably varied between 0.2 and 7.5 nkat g⁻¹ of juice, whereas PE activities were more uniform (0.4–1.5 PE units g⁻¹). In all juices, except ‘Ortanique’ juice, heating ≥42 °C for 12 s reduced POD activity below 4.3% of the maximum activity in fresh Citrus juice. Thermal tolerance of the thermo-labile PE fraction was overall much higher, but varied among juices during heating at temperatures ≤62 °C. Overall thermal resistance of PE was though comparable, with deactivation exceeding 84%, mostly even 90%, after thermal treatments ≥72 °C. Unlike POD, total PE activity proved to be an indicator of freshness that is universally applicable to Citrus juices derived from orange, mandarin, and lemon or blends thereof. Freshly squeezed juices can analytically be distinguished from cold-stored, minimally processed products that display an almost completely inactivated thermo-labile PE fraction and thus extended shelf life.     

Functional properties of different Korean sweet potato varieties

Eight different varieties of Korean sweet potatoes (SPs) were investigated to develop new healthy foods. The purple-fleshed SPs, ‘Shinjami’ and ‘Borami’, the orange-fleshed SPs, ‘Juwhangmi’ and ‘Shinwhangmi’, and the white/cream-fleshed dry-type SPs, ‘Shinyulmi’, ‘Shinchunmi’, ‘Yeonwhangmi’, and ‘Jeungmi’, were used. Alcohol insoluble solids (AIS), total dietary fiber (TDF), anthocyanin, carotenoid, and phenolic compounds contents for SP powders vary significantly (p<0.05) between different varieties. The TDF, anthocyanin, and the total phenolic compounds of SPs had the highest values in the purple-fleshed SPs (10.11–10.87%, 2.43–3.35 mg/g, and 454.13–638.79%, respectively) and the lowest values in the white/cream-fleshed dry-type SPs. The carotenoids of the orange-fleshed SPs were higher in ‘Juwhangmi’ than in ‘Shinwhangmi’. The color differences among the purple-fleshed SPs were 3–4 times larger than those of other SPs. The antioxidant activities of the purple-fleshed SPs were higher than those of other SPs.     

Application of magnetic immuno-polymerase chain reaction assay for detection of <Emphasis Type="Italic">Salmonella</Emphasis> spp. in chicken meats

Since contaminated chicken meats have been the principal foodborne source of the contamination of Salmonella to human beings and cultural detection methods are labor-intensive and time-consuming, a study evaluating the performance of the combination of two techniques that are immunomagnetic separation (IMS) and polymerase chain reaction (PCR) for the detection of Salmonella in chicken meats was conducted. The IMS and PCR assay combines selective extraction of Salmonella by specific antibodies with primer-specific (primer pair based on the sequence of invA gene) PCR amplification. Initially chicken meat samples, in which no Salmonella contamination had been determined by using ISO 6579 reference method, were inoculated with Salmonella Enteritidis culture and subsequently the shortest non-selective pre-enrichment time, that had been needed for the detection of approximately 1 or 10 CFU/mL chicken meat levels of target bacteria by magnetic immuno-PCR assay, was found by using 14, 12, 10 and 8-h periods. In conclusion, it was found that magnetic immuno-PCR assay was able to detect 1–10 CFU Salmonella/25 g chicken meat, after only incorporating a non-selective pre-enrichment period of 12 h. Therefore, an overall 16-h (magnetic immuno-PCR assay in conjunction with 12-h non-selective pre-enrichment) magnetic immuno-PCR assay statistically evaluated as sufficient (p = 0.182 > 0.05) for rapid and sensitive detection of approximately 1–10 CFU Salmonella from 25 g chicken meat samples. Accordingly, 16-h magnetic immuno-PCR assay can be promising for routine use in the detection of Salmonella in chicken meat samples, and it consequently may prevent the risk of Salmonella infections in regard to chicken meats.