Detection of the thaumatin-like protein gene from Brassica rapa var. oleifera in honey with the Proofman-LMTIA method

ABSTRACT

Honey adulteration has been on the rise with the increasing demand for high-quality honey. The ladders-shape melting temperature isothermal amplification (LMTIA) was a newly developed nucleic acid amplification technique, and the aim of this study was to establish a method combining the LMTIA with the proofreading enzyme-mediated probe cleavage (Proof-man probe) for detecting the thaumatin-like protein (TLP) genes of Brassica rapa var. oleifera in honey. The LMTIA primers was selected as a target to be designed, the temperature of the Proof-man LMTIA reaction was optimized, and the specificity, sensitivity, and applicability to the honey samples were determined. The results showed that the Proof-man LMTIA method was highly specific with a detection sensitivity of 30-40 copies genomic DNA/reaction at the optimal temperature of 60°C, and the amplification could be completed within 20 min. This method can detect the adulteration of low-price B. rapa honey into high-price honey.

KEYWORDS:

• Nucleic acid amplification techniques
• DNA probes
• honey
• Brassica rapa
• rapid tests

Acknowledgements

The authors wish to thank all members from the Key Laboratory of Biomarker Based Rapid-detection Technology for Food Safety of Henan Province, China, and Dr. Yanhong Liu from US Department of Agriculture for critical reading of the manuscript.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

The data that support this study are available from the corresponding author upon reasonable request.

Supplementary material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/19476337.2024.2351912

Additional information

Funding

This work was supported by the Natural Science Foundation of China [Grant No. 32172300], the Technology Project of Henan Province [Grant No. 212102310912] as well as Henan Province University Science and Technology Innovation Talent Project [Grant No. 23HASTIT048].