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Rapid and sensitive identification of the herbal tea ingredient Taraxacum formosanum using loop-mediated isothermal amplification.

Lai GH, Chao J, Lin MK, Chang WT, Peng WH, Sun FC, Lee MS, Lee MS - Int J Mol Sci (2015)

Bottom Line: Taraxacum formosanum (TF) is a medicinal plant used as an important component of health drinks in Taiwan.In this study, a rapid, sensitive and specific loop-mediated isothermal amplification (LAMP) assay for authenticating TF was established.In conclusion, using this novel approach, TF and its misused plant samples obtained from herbal tea markets were easily identified and discriminated by LAMP assay for quality control.

View Article: PubMed Central - PubMed

Affiliation: Graduate Institute of Biotechnology, National Chung Hsing University, Taichung 40402, Taiwan. just_dfm@hotmail.com.

ABSTRACT
Taraxacum formosanum (TF) is a medicinal plant used as an important component of health drinks in Taiwan. In this study, a rapid, sensitive and specific loop-mediated isothermal amplification (LAMP) assay for authenticating TF was established. A set of four specific LAMP primers was designed based on the nucleotide sequence of the internal transcribed spacer 2 (ITS2) nuclear ribosomal DNA (nrDNA) of TF. LAMP amplicons were successfully amplified and detected when purified genomic DNA of TF was added in the LAMP reaction under isothermal condition (65 °C) within 45 min. These specific LAMP primers have high specificity and can accurately discriminate Taraxacum formosanum from other adulterant plants; 1 pg of genomic DNA was determined to be the detection limit of the LAMP assay. In conclusion, using this novel approach, TF and its misused plant samples obtained from herbal tea markets were easily identified and discriminated by LAMP assay for quality control.

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Related in: MedlinePlus

Identification of Taraxacum formosanum contaminated with various ratios of adulterant’s genomic DNA using the LAMP method. The genomic DNA of T. formosanum and non-target DNA (genomic DNA of Ixeris chinensis) were mixed in the ratio 1:10 (lane 4), 1:50 (lane 3) and 1:100 (lane 2), respectively. Lane M, DNA marker; lane N, negative control (genomic DNA of Ixeris chinensis); lane 1, 130 ng genomic DNA of each species, Taraxacum officinale, Ixeridium laevigatum, Youngia japonica, Ixeris chinensis and Emilia sonchifolia var. javanica, were all mixed with 130 ng genomic DNA of T. formosanum into one sample.
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ijms-16-01562-f005: Identification of Taraxacum formosanum contaminated with various ratios of adulterant’s genomic DNA using the LAMP method. The genomic DNA of T. formosanum and non-target DNA (genomic DNA of Ixeris chinensis) were mixed in the ratio 1:10 (lane 4), 1:50 (lane 3) and 1:100 (lane 2), respectively. Lane M, DNA marker; lane N, negative control (genomic DNA of Ixeris chinensis); lane 1, 130 ng genomic DNA of each species, Taraxacum officinale, Ixeridium laevigatum, Youngia japonica, Ixeris chinensis and Emilia sonchifolia var. javanica, were all mixed with 130 ng genomic DNA of T. formosanum into one sample.

Mentions: To identify the T. formosanum within a mixed sample, multiple adulterant plants mixed into one sample were used as a contamination test. The genomic DNA of T. formosanum and non-target DNA (genomic DNA of Ixeris chinensis) were mixed in the ratio 1:100, 1:50 and 1:10 (where “1” represents 1 ng), respectively. After LAMP amplification, positive results were detected in all three of the different ratios of mixed samples (Figure 5, lane 2 to lane 4). These DNA diluents demonstrated that the DNA of T. formosanum may be amplified using TF LAMP primers when an increase from 10 to 100 ng of non-target DNA was added in reaction mixtures. Additionally, when 130 ng genomic DNA of Taraxacum officinale, Ixeridium laevigatum, Youngia japonica, Ixeris chinensis and Emilia sonchifolia var. javanica were mixed with 130 ng genomic DNA of T. formosanum into one sample, the LAMP product of T. formosanum was amplified using TF LAMP primers (Figure 5, lane 1). As a result, the developed LAMP method for TF identification was found to be selective and reliable also when DNA of T. formosanum is contaminated with genetic material of an adulterant plant.


Rapid and sensitive identification of the herbal tea ingredient Taraxacum formosanum using loop-mediated isothermal amplification.

Lai GH, Chao J, Lin MK, Chang WT, Peng WH, Sun FC, Lee MS, Lee MS - Int J Mol Sci (2015)

Identification of Taraxacum formosanum contaminated with various ratios of adulterant’s genomic DNA using the LAMP method. The genomic DNA of T. formosanum and non-target DNA (genomic DNA of Ixeris chinensis) were mixed in the ratio 1:10 (lane 4), 1:50 (lane 3) and 1:100 (lane 2), respectively. Lane M, DNA marker; lane N, negative control (genomic DNA of Ixeris chinensis); lane 1, 130 ng genomic DNA of each species, Taraxacum officinale, Ixeridium laevigatum, Youngia japonica, Ixeris chinensis and Emilia sonchifolia var. javanica, were all mixed with 130 ng genomic DNA of T. formosanum into one sample.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4307320&req=5

ijms-16-01562-f005: Identification of Taraxacum formosanum contaminated with various ratios of adulterant’s genomic DNA using the LAMP method. The genomic DNA of T. formosanum and non-target DNA (genomic DNA of Ixeris chinensis) were mixed in the ratio 1:10 (lane 4), 1:50 (lane 3) and 1:100 (lane 2), respectively. Lane M, DNA marker; lane N, negative control (genomic DNA of Ixeris chinensis); lane 1, 130 ng genomic DNA of each species, Taraxacum officinale, Ixeridium laevigatum, Youngia japonica, Ixeris chinensis and Emilia sonchifolia var. javanica, were all mixed with 130 ng genomic DNA of T. formosanum into one sample.
Mentions: To identify the T. formosanum within a mixed sample, multiple adulterant plants mixed into one sample were used as a contamination test. The genomic DNA of T. formosanum and non-target DNA (genomic DNA of Ixeris chinensis) were mixed in the ratio 1:100, 1:50 and 1:10 (where “1” represents 1 ng), respectively. After LAMP amplification, positive results were detected in all three of the different ratios of mixed samples (Figure 5, lane 2 to lane 4). These DNA diluents demonstrated that the DNA of T. formosanum may be amplified using TF LAMP primers when an increase from 10 to 100 ng of non-target DNA was added in reaction mixtures. Additionally, when 130 ng genomic DNA of Taraxacum officinale, Ixeridium laevigatum, Youngia japonica, Ixeris chinensis and Emilia sonchifolia var. javanica were mixed with 130 ng genomic DNA of T. formosanum into one sample, the LAMP product of T. formosanum was amplified using TF LAMP primers (Figure 5, lane 1). As a result, the developed LAMP method for TF identification was found to be selective and reliable also when DNA of T. formosanum is contaminated with genetic material of an adulterant plant.

Bottom Line: Taraxacum formosanum (TF) is a medicinal plant used as an important component of health drinks in Taiwan.In this study, a rapid, sensitive and specific loop-mediated isothermal amplification (LAMP) assay for authenticating TF was established.In conclusion, using this novel approach, TF and its misused plant samples obtained from herbal tea markets were easily identified and discriminated by LAMP assay for quality control.

View Article: PubMed Central - PubMed

Affiliation: Graduate Institute of Biotechnology, National Chung Hsing University, Taichung 40402, Taiwan. just_dfm@hotmail.com.

ABSTRACT
Taraxacum formosanum (TF) is a medicinal plant used as an important component of health drinks in Taiwan. In this study, a rapid, sensitive and specific loop-mediated isothermal amplification (LAMP) assay for authenticating TF was established. A set of four specific LAMP primers was designed based on the nucleotide sequence of the internal transcribed spacer 2 (ITS2) nuclear ribosomal DNA (nrDNA) of TF. LAMP amplicons were successfully amplified and detected when purified genomic DNA of TF was added in the LAMP reaction under isothermal condition (65 °C) within 45 min. These specific LAMP primers have high specificity and can accurately discriminate Taraxacum formosanum from other adulterant plants; 1 pg of genomic DNA was determined to be the detection limit of the LAMP assay. In conclusion, using this novel approach, TF and its misused plant samples obtained from herbal tea markets were easily identified and discriminated by LAMP assay for quality control.

Show MeSH
Related in: MedlinePlus