PCR (Polymerase Chain Reaction) and fluorescent PCR rapid detection method of bee sacbrood diseases
A technology for cystic larvae and detection methods, applied in fluorescence/phosphorescence, biochemical equipment and methods, and microbial determination/inspection, etc., can solve the problems of low sensitivity and specificity, reduce drug residues, and facilitate detection method, control effect
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[0029] The steps of the present invention are:
[0030] a, first collect adult bee specimens and larval specimens;
[0031] b. Primer design and synthesis:
[0032] Primers for PCR detection of bee cystic larvae:
[0033] SBV 1 5’-ACC AAC CGA TTC CTC AGT AG-3’
[0034] SBV 2 5'-CCT TGG AAC TCT GCT GTG TA-3'
[0035] Taqman probes and primers for real-time fluorescent PCR detection of bee cystic larvae:
[0036] SBV311F 5'-AAGTTGGAGGCGCGTAATTG-3'
[0037] SBV380R 5'AAATGTCTTCTTACTAGAGGTAAGGATTG-3'
[0038] SBV331T 5’-FAM-CGGAGTGGAAAGATTATCTACAATCCTTACCTCTA-TAMRA-3’
[0039] c. (1) Take a single bee, wash it with PBS solution for several times, discard as much excess PBS as possible, add 500 μL Trizol reagent, grind as much as possible with a grinder, then add 500 μL Trizol reagent, mix quickly, and stand at room temperature for 10 minutes ;
[0040] (2) Add 200 μL of chloroform, close the lid tightly, shake vigorously by hand for 15 seconds, and let stand at room ...
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