Kit and method for identifying and detecting Cordyceps sinensis

Abstract

The invention provides a method for identifying and detecting Cordyceps sinensis, wherein the method comprises steps of: (a) extracting ribosome DNA from a sample to be detected; (b) applying DNA extracted in step (a) as a template to perform real time polymerase chain reaction (PCR) using a primer pair shown by SEQ ID NO.1 and SEQ ID NO.2 and a probe shown by SEQ ID NO.3 to obtain amplification products; and (c) determining the authenticity and contents of the Cordyceps sinensis in dependence on the existence and intensity of the fluorescences of the amplification products. The invention also provides a kit which comprises the primer pair and the probe. The invention can qualitatively identify the authenticity of the Cordyceps sinensis and detect the Cordyceps sinensis contents of a sample with a lowest detection limitation of 0.0001 ng / ul, thus the invention is more sensitive than prior arts; and the invention can also effectively detect the Cordyceps sinensis samples in which nucleic acid is severely degraded after processing, and detecting results are more accurate and more effective than those obtained using prior arts.

Description

technical field [0001] The invention relates to a method for qualitatively and quantitatively detecting Cordyceps sinensis and a kit used in the method. Background technique [0002] Cordyceps sinensis is a complex of larvae and larvae produced by Cordyceps sinensis (Berk.) Saccardo (1878) of the family Ergotaceae parasitizing on the larvae of Lepidoptera, Batmothidae, and Hepialus. In the market of Chinese medicinal materials, the asexual mycelium cultured by its artificial fermentation and the related species similar to Cordyceps sinensis (such as Cordyceps japonica (also known as Cordyceps Hawkes) C. hawkesii Gray, Cordyceps daishii C. taii Z.Q.Liang & A.Y.Liu, Gunni Cordyceps C.gunnii (Berk.) Berk., Liangshan Cordyceps C.liangshanensis Zhang & Liu) and other counterfeit and shoddy products flooded the market, and even ground silkworms, white silkworms, and molded "Cordyceps" appeared Such fake and inferior products have caused confusion in the market of Cordyceps sinens...

AI Technical Summary

Problems solved by technology

However, this method has the following limitations: (1) After PCR amplification, DNA sequencing (needs 2-3 days) or gel electrophoresis (about 1.5 hours) must be performed to determine the results, and the results cannot be determined after PCR amplification. Therefore, the detection speed and efficiency are subject to certain restrictions; (2) This method is a qualitative detection method, that is, it can only detect whether Cordyceps sinensis is "present" or "absent", and cannot be used in technical (3) Because the spacer sequence of Cordyceps sinensis rDNA is short and the primer design is difficult, the sequence length of ordinary PCR amplification and identification is generally about 300bp. It may be severely degraded to less than 300bp. At this time, the amplification of the DNA chain cannot be achieved by ordinary PCR technology, resulting in inaccurate detection results; (4) This method needs to further perform gel electrophoresis on conventional PCR products, and the process involves nucleic acid dyes, etc. The operation process of toxic substances is easy to cause pollution and has a certain degree of harm to the human body

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