Chinese Journal of Tissue Engineering Research ›› 2022, Vol. 26 ›› Issue (2): 171-175.doi: 10.12307/2022.029

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Identification and expression of exogenous target genes in transgenic tomato

Guan Weiwei1, 2, 3, Gu Yu2, Guan Xiaoyan2, 3, Wu Jiayuan2, 3, Bai Guohui3, Tian Yuan2, 3, Liu Jianguo2, 3   

  1. 1Haikou Affiliated Hospital of Central South University Xiangya School of Medicine · Hainan Stomatological Center, Haikou 570208, Hainan Province, China; 2Stomatological Hospital Affiliated to Zunyi Medical University, Zunyi 563099, Guizhou Province, China; 3Key Laboratory of Oral Disease Research in Guizhou Higher Education Institutions · Zunyi Key Laboratory of Oral Disease Research, Zunyi 563006, Guizhou Province, China
  • Received:2020-11-27 Revised:2020-12-25 Accepted:2021-01-30 Online:2022-01-18 Published:2021-10-27
  • Contact: Liu Jianguo, MD, Professor, Chief physician, Doctoral supervisor, Stomatological Hospital Affiliated to Zunyi Medical University, Zunyi 563099, Guizhou Province, China; Key Laboratory of Oral Disease Research in Guizhou Higher Education Institutions · Zunyi Key Laboratory of Oral Disease Research, Zunyi 563006, Guizhou Province, China
  • About author:Guan Weiwei, Master, Associate chief physician, Haikou Affiliated Hospital of Central South University Xiangya School of Medicine · Hainan Stomatological Center, Haikou 570208, Hainan Province, China; Stomatological Hospital Affiliated to Zunyi Medical University, Zunyi 563099, Guizhou Province, China; Key Laboratory of Oral Disease Research in Guizhou Higher Education Institutions · Zunyi Key Laboratory of Oral Disease Research, Zunyi 563006, Guizhou Province, China
  • Supported by:
    Guizhou Provincial Party Committee Organization Department Talent Base Project, No. RCJD2019-9 (to LJG); Provincial and Municipal Science and Technology Cooperation Special Project, No. (2014)41 (to LJG); First-batch Talent Base Project of Zunyi Municipal Organization Department of Zunyi Municipal Party Committee, No. (2019)69 (to LJG); Hainan Provincial Health Industry Scientific Research Project, No. 20A200086 (to GWW)

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Abstract: BACKGROUND: Edible vaccines from transgenic plants is to integrate exogenous genes into the plant genome with plants as carriers, further activating animal or human immune system to obtain specific immunity. However, the continuous low expression of exogenous genes has been unable to achieve satisfactory immune effects.
OBJECTIVE: To detect the foreign fused gene pacA-ctxB gene and interest protein expression level in the transgenic tomato by molecular biological technique, providing a research basis for further observation of the anti-caries effect of edible caries vaccines. 
METHODS: After extracting the total DNA of transgenic tomato leaves, the exogenous fused gene pacA-ctxB was detected by PCR. There were three groups in the experiment: a positive control group (plasmid p2355-EPC10), a blank control group (an ordinary tomato plant, Hongkang 219), and a transgenic group (nine transgenic tomato plants). Total proteins were extracted and quantitatively tested with BCA kit and expression of PAcA/CTB in the transgenic tomato was analyzed by western blot and ELISA. There were two groups for detection of target proteins: a transgenic group (five transgenic tomato plants expressing exogenous chimeric genes (positive PCR test) and a blank control group (one ordinary tomato plant).
RESULTS AND CONCLUSION: PCR amplification results showed that there were 5 of the 9 strains of transgenic tomatoes in which the specific amplification bands were about 1.7 kb, accounting for 55% of the total detected transgenic plants. The total protein in the transgenic tomato was 3.15 g/L. Western blot results showed high density bands at 5.8 kD for PCR-positive transgenic tomato protein samples, and no specific bands were found in non-transgenic tomato protein samples. ELISA results showed that the expression of target proteins was about 4.12 mg/L, accounting for 0.13% of the total soluble protein in the transgenic tomato. To conclude, the exogenous fusion gene pacA-ctxB can be expressed in tomato plants and produce target proteins.

Key words: anti-caries vaccine, transgenic tomato, decayed tooth

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