Journal of Food Hygiene and Safety (한국식품위생안전성학회지)

The Korean Society of Food Hygiene and Safety (한국식품위생안전성학회)
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Development of Raw Material Identification Method of Changnan-jeot and Gaiyang-jeot Using Multiplex PCR and Real-Time PCR

Multiplex PCR과 Real-Time PCR을 이용한 창난젓과 가이양젓 원료 검사법 개발

  • Choi, Seong Seok (Basic Science Research Institute, Pukyong National University) ;
  • Seo, Yong Bae (Basic Science Research Institute, Pukyong National University) ;
  • Kim, Jong-Oh (Institute of Marine Biotechnology, Pukyong National University) ;
  • Yang, Ji-Young (Department of Food Science & Technology, Pukyoung National University) ;
  • Shin, Jiyoung (Institute of Food Science, Pukyong National University) ;
  • Kim, Gun-Do (Department of Microbiology, Pukyong National University)
  • Received : 2021.06.01
  • Accepted : 2021.08.09
  • Published : 2021.08.30
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Abstract

In this study, multiplex PCR and real-time PCR were performed on Theragra chalcogramma (walleye pollock), Pangasianodon hypophthalmus (iridescent shark) and their processed foods, such as changnan-jeot and gaiyang-jeot (salted iridescent shark intestine). Species-specific primers for T. chalcogramma and P. hypophthalmus were designed, and genomic DNA was directly extracted from each sample to perform single PCR and multiplex PCR. As a result of PCR, in the case of single PCR, PCR bands of T. chalcogramma (297 bp) and P. hypophthalmus (132 bp) were identified, and in the case of multiplex PCR, it was confirmed that amplification occurred without cross-reaction between T. chalcogramma and P. hypophthalmus. As a result of checking the PCR sensitivity, the concentration of genomic DNA was detected up to 0.1 ng/µL in both single PCR and multiplex PCR. The real-time PCR results showed that the average Ct value of T. chalcogramma was 20.765±0.691, and the average Ct value of P. hypophthalmus sample was 35.719±1.828 in the T. chalcogramma species-specific primers. In the P. hypophthalmus species-specific primers, the average Ct value of the T. chalcogramma sample was 35.996±1.423, and the mean Ct value of the P. hypophthalmus sample was 20.096±0.793. These results demonstrated the significant differences in the efficiency, specificity and cross-reactivity of species-specific primers in real-time PCR. Based on these findings, 7 of changnan-jeot or gaiyang-jeot products were confirmed by multiplex PCR and real-time PCR, and valid results were confirmed in all samples.

본 연구에서 multiplex PCR과 real-time PCR을 이용하여 창난젓의 원료를 감별할 수 있는 새로운 판별법을 개발하였다. 명태와 가이양의 종 특이 프라이머를 디자인하고, 명태와 가이양의 genomic DNA를 template로 single PCR과 multiplex PCR을 실시하였다. PCR을 실시한 결과, single PCR에서 명태(297 bp)와 가이양(132 bp)에 해당하는 PCR 밴드를 확인하였으며 교차 반응이 일어나지 않는 것을 확인하였다. Multiplex PCR에서 명태와 가이양 사이에 교차반응 없이 증폭이 일어나는 것을 확인하였다. Real-time PCR 결과, 명태 종 판별 프라이머에서 명태의 Ct 평균값은 20.765±0.691, 가이양 시료에서 Ct 평균값은 35.719±1.828이었으며, 가이양 종 판별 프라이머에서 명태 시료의 Ct 평균값은 35.996±1.423, 가이양 시료의 Ct 평균값은 20.096±0.793으로 프라이머의 효율성, 특이성 및 교차 반응성에서 유의한 차이가 나타났다. 이러한 결과를 바탕으로 시중에서 판매되는 7개 제품을 multiplex PCR 및 real-time PCR로 확인하였으며, 모든 시료에서 유효한 결과를 확인하였다. 본 연구에서 제작된 명태와 가이양에 대한 종 특이적 프라이머는 가공된 젓갈 시료의 원료의 판별 가능하며, 이러한 결과는 식품안전관리에 기여할 수 있을 것으로 기대된다.

Keywords

Acknowledgement

이 논문은 2021년 해양수산부 재원으로 해양수산과학기술진흥원의 지원을 받아 수행된 연구임(수산생물 원산지 판별기술 및 현장단속 키트개발)

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