Reagent box for simultaneously detecting diversified components of animal origin and application of reagent box
A technology of animal-derived ingredients and kits, which is applied in the direction of microbial determination/testing, biochemical equipment and methods, and can solve problems such as limitations in the scope of application
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Embodiment 1
[0053] Example 1 Screening of primers
[0054] Download the primer design template sequence from NCBI genebank, as shown in Table 4.
[0055] Table 4 Template sequence list
[0056] loci Serial Genebank number MCW248 G32016.1 FCA955 AY434939.1 PEZ1 AY672136.1 MAF33 M77200.1 cds BC081853.1 HLJ392 JN687127.1 SW742 AF235351.1 Sat7 X99888.1 CAUD056 AY493301.1 HMS3 X74632.1 BM2113 M97162.1 CFT073 AE014075.1
[0057] The sequence of each locus is compared in the genebank, primers are designed at both ends of the STR sequence, and the unique primers with better specificity are selected and compared by blast. After a large number of repeated tests, the DNA template of each species is amplified in the multiple amplification system. When designing primers, it is necessary to repeatedly debug in the primer design software to ensure that all primers are of moderate length, have similar physical...
Embodiment 2
[0074] Embodiment 2, actual sample test
[0075] Buy Your Own 12 Types of Meat on the Market, Using QIAamp ? The DNA Blood Mini Kit was used to extract DNA from 12 kinds of meat.
[0076] Detect the concentration of 12 extracted DNA with a spectrophotometer, and dilute to 0.5ng / μL as a template.
[0077] Add samples according to the sample loading system in Table 6, and perform amplification according to the amplification program in Table 3.
[0078] Table 6 PCR loading system
[0079] components volume Reaction Mix 10.0 μL DNA template 2μl primer mix 5 μL Hot start Taq enzyme (5U / μL) 0.5 μL wxya 2 o Make up to 25.0 μL
[0080] The Reaction Mix mentioned in it is MgCl 2 7.5mM, Tris-HCl buffer 125mM, KCl 125mM, dNTPs 7.5mM, BSA 2mg / ml.
[0081] The amplification results were analyzed according to the electrophoresis detection and data analysis methods in Example 1, and 3 tubes of amplification were repeated for each tem...
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