Method for detecting saiga tatarica horn ingredients in mixture and primers used in same
A mixture, antelope horn technology, applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc., can solve the problem of high subjectivity
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Embodiment 1
[0058] Example 1 Detection of Antelope Horn Components in Lingyang Qingfei Pills (Lingyangjiao Fang)
[0059] 1. Extraction of total DNA from antelope horns
[0060] This extraction step can be performed according to conventional operations in the field. The specific operation of this embodiment is as follows: antelope horns are ground into powder by liquid nitrogen, 200 μg is taken and 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 CW05...
Embodiment 2
[0092] Example 2 Identification of Antelope Horn Components in Niuhuang Qingxin Pills (Antelope Horn Prescription)
[0093] Select 3 batches (10 pills / batch) of Niuhuang Qingxin Pills (Antelope Horn Prescription) produced by Tongrentang—a total of twenty-nine medicines, of which the Antelope Horn component accounts for about 0.9% (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.
[0094] 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 antelope species-specific primers can effectively amplify the target fragment ...
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