Influence of Heat Processing on DNA Degradation and PCR-Based Detection of Wild-Type and Transgenic Maize

Abstract

Reliable detection of genetically modi ed (GM) maize is signi cant for food authenticity, labelling, quality, and safety assessment.is study aims to evaluate the factors in uencing degradation and polymerase chain reaction (PCR) ampli cation of DNA from the wild type and transgenic maize (events Bt-176 and MON810) during thermal treatment at 100°C and 121°C. A new PCR method was developed targeting the Cry1Ab gene to detect insect-resistant GM plants. e yield of genomic DNAs extracted by the DNeasy plant mini kit dramatically decreased while DNAs obtained by cetyltrimethyl ammonium bromide-(CTAB-) based method did not show any visible changes in the yield by the time of processing. Treatment at 100°C did not signi cantly a ect either genomic DNAs or amplicons. Heating at 121°C induced time-dependent degradation of genomic DNAs and exogenous Cry1Ab gene; however, it did not have any considerable in uence on the exogenous 141 bp amplicons or endogenous amplicons in the range of 102 bp to 226 bp with the exception of the event MON810 extracted by the DNeasy plant mini kit. More yield was observed at 226 bp than 140 bp fragment of the invertase gene. e 141 bp fragment of the transgenic CaMV 35S promoter exhibited the highest thermal stability of all the examined amplicons. Analysis of foodstu s demonstrated 102 bp amplicons speci c for the zein gene as the e ective marker to detect maize in the processed foods. e obtained results demonstrate that PCR-based detection of the wild type and transgenic maize is dependent on the combination of di erent parameters of crucial factors such as temperature and duration of exposure, transgenic event, DNA extraction method, DNA marker, and size and location of amplicons.

RNase A Enzyme Modification of Optimized SDS Protocol for DNA Extraction Suitable for Real-Time PCR Screening of GMOs

“…1B) even though the smear was less blurred than in the protocol without RNase A enzyme (12). Thermal processing could affect DNA integrity and stability, causing fragmentation of the genomic DNA (18).…”

Sensitivitas Real-Time Polymerase Chain Reaction dengan Primer Tanabe dalam Mendeteksi Gelatin Babi pada Confectionery

“…Hasil uji penentuan nilai LOD gelatin babi (% b/b) di dalam produk confectionery Table 2 Pemanasan dapat menyebabkan DNA mengalami fragmentasi yang dapat memengaruhi hasil analisis real-time PCR (Mano et al, 2017). Bitskinashvili et al (2019) menyatakan bahwa salah satu faktor kritis yang mempengaruhi analisis realtime PCR dalam identifikasi ingredien pangan adalah waktu kontak pemanasan. Pada penelitian tersebut dijelaskan bahwa DNA mengalami degradasi pada saat sampel dipanaskan pada suhu 121°C, yang intensitasnya tergantung pada waktu pemanasan.…”