Establishment and evaluation of a rapid PCR-colloidal gold test strip method for the detection of Fritillaria ussuriensis
This study design of specific identification primers for the ITS2 sequence of F.ussuriensis.The reaction system and conditions were optimized,and PCR-nucleic acid test strips were constructed to realize the visual detection of F.ussuriensis Bark.Through molecular cloning and sequencing technology,we constructed a positive control for F.ussuriensis DNA and formulated quality standards.The established method was evaluated for sensitivity,specificity and reproducibility,and the authenticity of the commercially available samples was identified.Results demonstrated that based on the ITS2 sequences,F.ussuriensis and its mixed forgeries could be distinguished.The PCR products of the authentic F.ussuriensis on test strips showed two bands in the T and C lines,while the pseudo products and negative control showed only one band in the C line,which was consistent with the results of agarose gel electrophoresis.The specificity was 100％,and the sensitivity of the PCR-nucleic acid test strip was up to 0.1 ng·μL-1,which was 10 times higher than that of the gel electrophoresis assay.11 out of 16 commercially available samples of F ussuriensis were qualified,and 5 were unqualified.Collectively,the PCR-nucleic acid test strip method established in this study is specific,rapid,accurate and visualized,which can provide a new technical idea for the detection of F.ussuriensis.
F.ussuriensismolecular cloningITS2 sequencenucleic acid test stripvisualization
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