Method for distinguishing astragalus root from its counterfeit drug by using SSR molecular marking process
A labeling method, the technology of Radix Astragali, applied in the direction of biochemical equipment and methods, pharmaceutical formulations, microbial determination/inspection, etc.
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0021] Embodiment 1, CTAB extracts the total DNA of dried Radix Astragali and its counterfeit products (hollyhock, upland cotton, alfalfa, round leaf mallow, sweet clover, mallow, golden pheasant)
[0022] (1) Add liquid nitrogen to the fresh leaves and grind them quickly, and the dry materials are crushed with a pulverizer before, and the powder is taken.
[0023] (2) Add preheated 65°C CTAB extraction buffer (pH7.8) to each tube
[0024] 700mmol / L NaCl,
[0025] 50mmol / L Tris-HCl,
[0026] 10mmol / L EDTA,
[0027] 1% CTAB,
[0028] 1% mercaptoethanol,
[0029] Mix well, and incubate at 65°C for about 30-60 minutes.
[0030] (3) Take it out and centrifuge at 10,000 rpm for 10 minutes.
[0031] (4) Take the supernatant and centrifuge at 10,000 rpm for 10 minutes, and keep the supernatant.
[0032] (5) Add an equal volume of chloroform-isoamyl alcohol (24:1), mix well, and rotate at 10,000 rpm for 10 minutes.
[0033] (6) Add 0.1 volume of 2M ammonium acetate to the supe...
Embodiment 2
[0040] Embodiment two, PCR amplification
[0041] number 1
chain of justice
5′CAAAACATAAAAAAGGTGAGA 3′
antisense strand
5′AAGAACCACACTAATATTATT 3′
number 2
chain of justice
5′GGAAGAAAGTATTGGTCTGT 3′
antisense strand
5′AGGAGAGAGTGGAGAGATTA 3′
number 3
chain of justice
5′ TTCGAAGCATCCAAGG 3′
antisense strand
5′AAAAGACAAAACATACTATAAAA 3′
number 4
chain of justice
5′CGTGCCAAATTACATCA 3′
antisense strand
5′TGATGGGAACAAGTACATAA 3′
number 5
chain of justice
5′ ATGGTTTTTTTCCTGCCCTTT 3′
antisense strand
5′AACAAGCCCCCCTAAAAGAA 3′
number 6
chain of justice
5′ CTTTGGCGCTGACACAT 3′
antisense strand
5′CCCAGTGAACATTTTAGGATCA 3′
number 7
chain of justice
5′AAGCTTAGAAGCAAACCAAAGAT 3′
antisense strand
5′ TGTTTTTCATTTTCACCCCAAG 3′
number 8
chain of justice
5′AATGAGGAACCTTTGACCCC 3′
...
Embodiment 3
[0064] Embodiment three, SSR is to Radix Astragali and its counterfeit identification result
[0065] The results of electrophoresis were converted into patterns of electrophoresis bands using the biological software Orthogonality-Experiment Assistant (version: 2.0) (Figs. 8-13). The site with a band was counted as 1, and the site without a band was counted as 0, and the results in the negative control were subtracted from it. And calculate the similarity index Sxy between each material for each pair of primers according to the following formula:
[0066] S xy =2×N xy / N x +N y
[0067] N x is the number of alleles of material X, N y is the number of alleles of material X, N xy is the number of alleles shared by X and Y.
[0068] Calculate the PIC value (polymorphism index content) according to the following formula:
[0069] PIC = 1 - Σ n = 1 ...
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More 