(Size: 100 rxns × 20 µl Systems)
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Abbexa's Animal Tissue PCR Kit uses a unique lysis buffer to lyse fresh or frozen animal tissues and blood. The resulting lysate without purification can be directly used as template for PCR amplification. Kit Contents:
| Component || 100 rxns × 20 µl Systems || 500 rxns × 20 µl Systems|
|AD1 Buffer||4 ml ||20 ml|
|AD2 Buffer ||1 ml ||5 ml|
|AD3 Buffer ||4 ml ||2 × 10 ml|
|2X PCR SuperMix (with dye) ||1 ml ||5 × 1 ml|
|Nuclease-Free Water ||5 ml ||25 ml|
Research Articles on Animal Tissue PCR Kit
||Animal Tissue PCR Kit
||Store at -20 °C for up to two years. Avoid repeated freeze/thaw cycles.
||PCR SuperMix (with dye) contains ≤ 30% glycerol. The formulations of the AD1, AD2 and AD3 Buffers are proprietary.
|Directions for use
||Genomic DNA Extraction:
- Immediately prior to carrying out the assay, prepare a AD1/AD2 Buffer mixture solution at a ratio of 4:1 (e.g. 40 µl of AD1 Buffer and 10 µl of AD2 Buffer) and mix thoroughly. 50 µl of AD1/AD2 Buffer mixture solution is sufficient for 1 sample/reaction. This mixture can be stored for up to 2 hours at room temperature.
- Prepare samples as follows:
- Mammalian cells - Recommended assay amount: ≤ 106 cells
Pellet cells by centrifugation and remove the supernatant. Add 50 µl of AD1/AD2 Buffer mixture solution to the pellet and mix thoroughly by pipetting up and down.
- Saliva or Blood - Recommended assay amount: ≤ 10 µl
Add saliva or blood to 50 µl of AD1/AD2 Buffer mixture solution and mix thoroughly by pipetting up and down.
- Animal tissues - Recommended assay amount: ≤ 10 mg (Mouse tail: ≤ 0.5 cm; Mouse ear: ≤ 0.5 cm2)
Cut tissue into small pieces using a sterile blade or scissors. Add 50 µl of AD1/AD2 Buffer mixture solution and mix thoroughly by pipetting up and down.
- Allow to stand at room temperature for 10 minutes (for samples that are hard to lyse such as hair tissue, the sample mixture can instead be incubated at 55 °C for 10 minutes), then incubate at 95 °C for 3 minutes.
- Add 40 µl of AD3 Buffer, then mix thoroughly. The lysate can be used as a PCR template, stored at 2-8 °C for up to 3 months, or stored at -20 °C for up to 6 months.
Thermal Cycling Conditions:
| Component || Volume || Final Concentration |
| Unpurified Lysate || Variable (≤ 4 µl) || as required |
| Forward Primer (10 µM) || 0.4 µl || 0.2 µM |
| Reverse Primer (10 µM) || 0.4 µl || 0.2 µM |
|2X PCR SuperMix (with dye) || 10 µl || 1X|
| Nuclease-Free Water || Variable ||N/A |
| Total Volume || 20 µl ||N/A |
| Number of Cycles || Temperature || Time per Cycle|
| 1 cycle|| 94 °C || 5-10 min|
|30-35 cycles || 94 °C ||30 seconds|
|50-60 °C ||30 seconds|
|72 °C|| 1-2 kb/min|
|1 cycle ||72 °C||5-10 min|
- Ensure that the sample and reagents have completely thawed and are mixed well prior to use in the assay.
- If faint bands are observed, increase the quantity of PCR template used or increase the number of PCR cycles (maximum 40 cycles).
- If non-specific amplification bands are observed, adjust the annealing temperature or reduce the quantity of PCR template used.
||Shipped within 10-20 working days.
||This product is for research use only.