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Early detection of the zearalenone (ZEA) chemotype of Fusarium species could be a precautionary measure for preventing ZEA contamination in rice. In this study, a multiplex polymerase chain reaction (mPCR) assay for detecting ZEA-producing fungi in rice was established using a set of four primers targeting the ZEA biosynthesis genes PKS3, PKS13, ZEB1, and ZEB2. Two mPCR approaches were used: one that amplified the DNA obtained from Fusarium isolates (conventional method) and another that directly amplified the target DNA from rice samples without time-consuming DNA isolation (direct method). The two mPCR methods showed high sensitivity in detecting ZEA-producing species, with a detection limit of 1.25 pg/μL of genomic DNA and 102 and 103 spores/g of white and brown rice, respectively. Both methods were specific for ZEA-producing species and gave four band patterns. The application of the two mPCR methods to 51 Fusarium isolates and 41 rice samples revealed that 31% (16 of 51) and 24% (10 of 41) of the samples were contaminated with ZEA-producing species, respectively. The mPCR results were further evaluated using high-performance liquid chromatography; in general, the two methods yielded similar results. These findings indicate that both mPCR methods are suitable for the detection of ZEA-producing Fusarium species in white and brown rice; however, the direct method yielded more rapid results.
This article was published in the following journal.
Name: International journal of food microbiology
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Methods for using more than one primer set in a polymerase chain reaction to amplify more than one segment of the target DNA sequence in a single reaction.
Methods used for detecting the amplified DNA products from the polymerase chain reaction as they accumulate instead of at the end of the reaction.
A heat stable DNA-DIRECTED DNA POLYMERASE from the bacteria Thermus aquaticus. It is widely used for the amplification of genes through the process of POLYMERASE CHAIN REACTION. EC 2.7.7.-.
In vitro method for producing large amounts of specific DNA or RNA fragments of defined length and sequence from small amounts of short oligonucleotide flanking sequences (primers). The essential steps include thermal denaturation of the double-stranded target molecules, annealing of the primers to their complementary sequences, and extension of the annealed primers by enzymatic synthesis with DNA polymerase. The reaction is efficient, specific, and extremely sensitive. Uses for the reaction include disease diagnosis, detection of difficult-to-isolate pathogens, mutation analysis, genetic testing, DNA sequencing, and analyzing evolutionary relationships.
A technique that labels specific sequences in whole chromosomes by in situ DNA chain elongation or PCR (polymerase chain reaction).
Polymerase Chain Reaction (PCR)
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Immunoassay - ELISA
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Food is any substance consumed to provide nutritional support for the body. It is usually of plant or animal origin, and contains essential nutrients, such as carbohydrates, fats, proteins, vitamins, or minerals. The substance is ingested by an organism ...