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Method for detecting capra hircus horn ingredients in mixture and primers used in same

A technology of mixtures and goat horns, applied in biochemical equipment and methods, measurement/testing of microorganisms, DNA/RNA fragments, etc., can solve problems of high subjectivity

Active Publication Date: 2012-04-25
DOJINDO +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 goat horns, 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 capra hircus horn ingredients in mixture and primers used in same
  • Method for detecting capra hircus horn ingredients in mixture and primers used in same
  • Method for detecting capra hircus horn ingredients in mixture and primers used in same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Example 1 Detection of Goat Horn Components in Lingyang Qingfei Pills (Goat Horn Recipe)

[0059] 1. Extraction of goat horn total DNA

[0060] This extraction step can be performed according to conventional operations in the field. The specific operation of this embodiment is as follows: goat horns are ground into powder by liquid nitrogen, and 200 μg is added to 1 ml of lysate B (10 mM Tris-HCl pH8.0, 0.1M EDTA pH8.0, 0.5% SDS, 200 μg / ml proteinase K, 0.4M DTT), digested at 55°C overnight, digested samples were boiled at 100°C for 10min, ice-bathed for 5min, centrifuged at 12000rpm for 15min, and the supernatant was extracted with phenolform. Then precipitate the DNA with 2 times the volume of absolute ethanol, wash the precipitate twice with 75% ethanol, and dissolve in an appropriate amount of double distilled water. The sample DNA was purified by a PCR purification kit (Kangwei Century PCR Product Purification Kit 50T, product number CW0521, and the specific oper...

Embodiment 2

[0093] Identification of Goat Horn Components in Niuhuang Qingxin Pills (Goat Horn Prescription) in Example 2

[0094] Select 3 batches (10 pills / batch) of Niuhuang Qingxin Pills (Goat’s Horn Prescription) produced by Tongrentang—a total of 29 medicines, of which the Goat’s Horn component accounts for about 9% (mass percentage) as the sample to be tested, extract The total DNA was subjected to single-plex PCR amplification with the primer pairs of SEQ ID Nos.1 and 2, SEQ ID Nos.3 and 4, and SEQ ID Nos.5 and 6, respectively. The extraction process of the total DNA of the sample to be tested, the PCR amplification and the analysis process of the amplification result are the same as in Example 1.

[0095] See results Figure 5 (Take the experimental results of one sample as an example, the rest are the same), using the DNA extraction method of the present invention and the designed goat species-specific primers can effectively amplify the target fragment from the corresponding f...

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Abstract

The invention provides a method for detecting capra hircus horn ingredients in a mixture and primers used in the same. The method mainly comprises the steps of extracting total DNA from the mixture to be detected; with the extracted total DNA as a template, utilizing specific primers to carry out PCR (polymerase chain reaction) amplification; and analyzing and judging the amplification results, wherein the specific primers are selected from one pair, two pairs or three pairs of such primers as SEQ ID Nos.1 and 2, SEQ ID Nos.3 and 4 and SEQ ID Nos.5 and 6. The invention further provides a kit which comprises the primers and is used for detecting capra hircus horn ingredients in the mixture, especially the crude medicines needing differentiating to judge whether the crude medicines are capra hircus horns or the traditional Chinese medicine preparations using capra hircus horns or substitutes. The primers have higher specificity and sensitivity.

Description

technical field [0001] The invention relates to a method for detecting goat horn components in mixtures, especially medicinal materials, using molecular biology techniques, in particular to a method for detecting goat horn components in medicinal materials, or traditional Chinese medicine preparations that use goat horn or substitutes for medicine, and the specific method used in the method. sex primers. Background technique [0002] Goat horn is the horn of bovine animals Qingyang and goat. It has analgesic and antipyretic effects similar to antelope horn. It can be used as a substitute for antelope horn. It is rich in resources, which is conducive to reducing costs and protecting rare animals. But at present, the traditional morphological method is mainly used for the identification of goat horns, which relies on the experience of the identification personnel and is highly subjective. Therefore, an accurate and effective identification method is needed. [0003] On the ot...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
Inventor 李志猛曹梦姚璐彭鹏张晓楠张丽娟
Owner DOJINDO
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