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Method for detecting Osteon Myospalacem Baileyi in mixture and primers used thereby

A technology of mixtures and sialons, applied in the fields of biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc., which can solve the problem of high subjectivity

Active Publication Date: 2012-04-25
BEIJING TONGRENTANG CO LTD +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, the traditional morphological method is mainly used for the identification of Theron bones, which relies on the experience of the identification personnel and is highly subjective. Therefore, an accurate and effective identification method is needed.

Method used

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  • Method for detecting Osteon Myospalacem Baileyi in mixture and primers used thereby
  • Method for detecting Osteon Myospalacem Baileyi in mixture and primers used thereby
  • Method for detecting Osteon Myospalacem Baileyi in mixture and primers used thereby

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Example 1 Identification of Sailong Bone Components in Zaizai Pills (Sailong Fang)

[0060] 1. Extraction of total DNA from Sialon bone

[0061]The extraction step can be performed in accordance with routine operations in the art. The specific operation of this example is as follows: Sialon bone is ground into small pieces with a mortar, and then decalcified by EDTA (10% EDTA solution (pH 7.0) for 48 hours before DNA extraction. 200 μg sample (Sialon after decalcification) Add 1 ml of Lysis Solution A (10 mM Tris-HCl pH8.0, 0.1 MEDTA pH8.0, 0.5% SDS, 200 μg / ml Proteinase K) to the bone, initial weight), digest at 55°C overnight. Centrifuge the digested samples at 12,000 rpm for 15 min to take the supernatant Carry out phenolic extraction. After that, use 2 times the volume of absolute ethanol to precipitate DNA, wash the precipitation twice with 75% ethanol, and dissolve in an appropriate amount of 50 μl of double distilled water. The sample DNA is purified by PCR kit ...

Embodiment 2

[0094] Example 2 Identification of the components of the sialon bone in the sialon rheumatism capsules

[0095] Select 3 batches (10 capsules / batch) of Sialon Rheumatism Capsules (one medicine in total) as the samples to be tested, and each sample takes 4g of capsule contents, extracts DNA, and uses the primer pairs SEQ ID Nos. 1 and 2, SEQ ID Nos. 3 and 4, and SEQ ID Nos. 5 and 6, respectively, were subjected to single-plex PCR amplification.

[0096] see the results Image 6 (Take the experimental result of one sample as an example, the rest are the same), the Sialon species-specific primer can amplify the target fragment from the corresponding patent medicine.

[0097] Image 6 Swimming lane 1 represents the detection result of the sample to be tested with Zaizaiwan (Leopard Bone); Swimming lane 2 represents the detection result of the sample to be tested with Sailong Fengshi Capsule (Leopard Bone); Swimming lane 3 represents the reconstruction of embodiment 1 Pills (Sai...

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Abstract

The invention provides a method for detecting Osteon Myospalacem Baileyi in a mixture and primers used thereby. The method mainly comprises the following steps: 1, extracting total DNA from a mixture to be detected; 2, carrying out PCR amplification through specific primers with the extracted total DNA as a template, wherein the specific primers are selected from one, two or three of SEQ ID Nos.1and SEQ ID Nos.2, SEQ ID Nos.3 and SEQ ID Nos.4, and SEQ ID Nos.5 and SEQ ID Nos.6; and 3, analyzing and determining amplification results. The invention further provides a kit which contains the primers and is used for discriminating whether a mixture to be detected is an Osteon Myospalacem Baileyi medicine or detecting Osteon Myospalacem Baileyi in a traditional Chinese medicinal preparation containing Osteon Myospalacem Baileyi or a substitute thereof. The primers of the invention have high specificity and sensitivity.

Description

technical field [0001] The present invention relates to a method for detecting sialon bone components in mixtures, especially medicinal materials, by applying molecular biology techniques, in particular to a method for detecting sialon bone components in medicinal materials, or traditional Chinese medicine preparations using sialon bones or substitutes for medicine, and the method specific primers used. Background technique [0002] Theron, scientific name plateau zokor (Myospalax Baileyi), mammal rodent family hamster, is a mammalian pest endemic to the Qinghai-Tibet Plateau. The dried whole bones of Sailong belong to the first class of new animal medicinal materials at the national level. However, the traditional morphological method is mainly used for the identification of sialon bones at present, which relies on the experience of the identification personnel and is highly subjective. Therefore, an accurate and effective identification method is required. [0003] On th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
Inventor 解素花姚璐曹梦杜菁王海刘名张薇张晓兰
Owner BEIJING TONGRENTANG CO LTD
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