Shrimp baculovirus (BP) nucleic acid detection kit (one-tube thermostat fluorescence method)
â—† Product Description
The animal disease detection series is based on a unique thermostatic fluorescence detection technology that can amplify specific nucleic acid fragments of diseases in food, animal tissues and the like, and judge the results by real-time amplification curve. This product is used for the detection of shrimp baculovirus disease (BP) with a detection limit of 10 3 copies/μl genomic DNA .
â—† Product composition (96 test)
031061LIII | |
Reagent | content |
A-BP-I | 22μL × 96 tubes |
BI | 90μL × 2 |
NG-I | 50μL × 3 |
PG-BP-I | 50μL × 2 |
â—† Applicable instruments
Dhelix-C, ESE Tube Scanner, Genie II, Deaou-308C and other constant temperature fluorescence detectors, ABI 7500, LightCycler480, CFX 96 and other fluorescent PCR instruments.
â—† Self-contained supplies and instruments
1 Sterilize 1.5mL or 2.0mL centrifuge tube; 2 ice box; 3 pipette (0.1-2.5μL, 0.5-10μL, 10-100μL, 100-1000μL) and matching sterilization tips; 4 centrifuge; 5 vortex Rotary mixer; 6 metal bath
â—† Notes
1. This reagent has high detection sensitivity. In order to prevent pollution, the experiment is to be partitioned.
1) First zone: sample preparation zone.
2) The second zone: the template addition zone.
3) Zone 3: Amplification and product analysis zone.
★ It is best to physically isolate the partitions to avoid pollution caused by human factors.
2. Work clothes and latex gloves are worn during the experiment, and tools are used independently in different areas. Gloves and lab coats need to be replaced.
3. Strictly follow the operation steps, reagent preparation and sample loading steps, please operate in strict accordance with the instructions on the ice box.
4. The components in the reaction solution are sensitive to light and should be stored away from light . The reagent should be completely thawed before use, but repeated freezing and thawing should be avoided. It is recommended to centrifuge for 30 seconds before use.
5. After the reaction is completed, the expansion tube should be placed in a sealed bag and discarded. On the same day, the lid is opened and the aerosol is easily contaminated. It is forbidden to open the lid.
6. Do not mix different batches of reagents used within the validity period.
â—† Sample processing
Samples prepared for treatment according to SN/T 1151.5-2015 Technical Specifications for Shrimp Baculovirus Quarantine or other standards are prepared for use.
For detailed steps, please follow the standard operation or check the food safety software.
â—† Experimental operation
1. Template preparation (sample preparation area)
It is recommended to use the aquatic animal virus genomic DNA/RNA extraction kit series. The specific process is detailed in the product manual.
2. Add a template (template add area, placed in the ice box)
The PCR tube containing the reaction solution was taken out, and the reagent was completely thawed. After thawing, the tube lid was opened and 1 μL of BI was added to the bottom of each tube, and the tube lid was centrifuged for 1 min. Open the tube cover and add 2 μL of template along each tube wall in the order of NG-I, sample template to be tested, and PG-BP-I. After the PCR tube cover was capped, it was centrifuged for 3 min, and the PCR amplification reaction was immediately performed.
3. Amplification reaction (amplification and product analysis area)
1 The constant temperature instrument was reacted at 63 ° C for 30 min.
2 If a real-time PCR instrument is used, the fluorescent group is selected as FAM, the quenching group is selected as None, 63 ° C for 15 s, 63 ° C for 45 s as a cycle, and the fluorescence signal is collected at 63 ° C for 45 s, 30 cycles.
For other instruments, please refer to the instrument manual for setting.
â—† Result judgment
1 The instrument automatically determines the result. If “positive†is displayed, the sample contains shrimp baculovirus disease (BP); if “negative†is displayed, the sample does not contain prawn baculovirus (BP) or the content is below the detection limit. . For example, if the curve rises between 20min and 30min and the automatic interpretation result is negative, it may be due to the low sample content, it is recommended to enrich the sample or extend the reaction time to 45min for review.
2 On the fluorescence quantitative PCR machine, the results were determined based on the presence or absence of the "S" type amplification curve. If there is an "S" type amplification curve, the sample contains shrimp baculovirus disease (BP); if there is no "S" type amplification curve, the sample does not contain shrimp baculovirus disease (BP) or the content is lower than Detection limit.
The result of the NG reaction tube showed "negative", and the result of the PG reaction tube showed "positive". The test result was valid, otherwise it was invalid. If the duplicate test results are still invalid, please contact technical support. 
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