Plant Tissue Direct PCR Kit (with Dye)
MCE Plant Tissue Direct PCR Kit (with Dye) is specifically designed for direct amplification from various plant leaf samples. The kit enables rapid release of genomic DNA from plant tissues (such as rice, maize, tobacco, rapeseed, etc.) in a single step. The resulting lysate can be directly used for downstream PCR amplification without the need for conventional DNA extraction and purification, thereby significantly simplifying the experimental workflow.
-
Storage :
Lysis Buffer: 4°C, 1 year. Neutralization Buffer, Direct PCR Master Mix (2×, with Dye): -20°C, 1 year. Avoid repeated freeze-thaw cycles.
- Manual (4931KB)
- COA
To place orders, for customer services and technical support, please contact: MedChemExpress USA
Tel: 609-228-6898 E-mail: [email protected] [email protected]
Description & Advantages
MCE Plant Tissue Direct PCR Kit (with Dye) is specifically designed for direct amplification from various plant leaf samples. The kit enables rapid release of genomic DNA from plant tissues (such as rice, maize, tobacco, rapeseed, etc.) in a single step. The resulting lysate can be directly used for downstream PCR amplification without the need for conventional DNA extraction and purification, thereby significantly simplifying the experimental workflow.
The kit is supplied with a ready-to-use PCR master mix containing all components required for amplification, including Taq DNA polymerase, dNTPs, an optimized buffer system, and tracking dyes. Only primers and template need to be added to initiate the PCR reaction. The amplified products can be directly analyzed by agarose gel electrophoresis without the addition of loading buffer, effectively reducing operational complexity and the risk of contamination. It is suitable for applications such as transgenic plant identification and plant genotyping.
Features of MCE Plant Tissue Direct PCR Kit (with Dye):
1. Low sample requirement: Efficient lysis can be achieved using only a small amount of plant tissue (as little as ~1 mm of leaf tissue).
2. High Specificity and Efficiency: Optimized Taq polymerase system with strong template affinity and high amplification specificity, ideal for genotyping and transgenic analysis.
3. User-Friendly Operation: The master mix contains tracking dyes, allowing PCR products to be directly loaded for electrophoresis.
Protocol
General Protocol
1. Plant leaf sample preparation
Method 1: Grinding lysis
1) Grinder method: Place a leaf disc approximately 5 mm in diameter in 50 μL Lysis Buffer, add two steel beads (about 3 mm), and grind at 45 Hz for 1 min. After disruption, the solution becomes green. Briefly centrifuge, keep the supernatant at 4°C, and use 1 μL for PCR.
2) Pipette-tip mashing: Young leaves are recommended. Place a leaf disc approximately 5 mm in diameter in 50 μL Lysis Buffer and mash thoroughly with a pipette tip until the solution becomes green. Briefly centrifuge, keep the supernatant at 4°C, and use 1 μL for PCR.
Method 2: Thermal lysis
Place a young leaf disc approximately 5 mm in diameter in 50 μL Lysis Buffer, ensure complete immersion, and heat at 95°C for 5-10 min. For difficult mature leaves, extend to 10-20 min. Mix, briefly centrifuge, and use the supernatant for PCR.
Method 3: Direct leaf amplification
Use a punch or blade to obtain an approximately 1 mm leaf disc and place it directly into the PCR mixture. For complex samples or long-fragment amplification, use a leaf piece smaller than 1 mm.
Note: a. Fresh young leaves are recommended; avoid the main vein when mature leaves are used;
b. For improved storage stability, mix lysate with Neutralization Buffer at a 5 : 1 ratio;
c. Mix and centrifuge processed leaf lysates and use only the supernatant for PCR, avoiding tissue debris.
2. PCR reaction system
| Component | 20 μL reaction | 50 μL reaction | Final concentration |
|---|---|---|---|
| Direct PCR Master Mix (2×, with Dye) | 10 μL | 25 μL | 1× |
| Forward Primer (10 μM) | 0.5 μL | 1 μL | 0.2-0.25 μM |
| Reverse Primer (10 μM) | 0.5 μL | 1 μL | 0.2-0.25 μM |
| Lysate supernatant (DNA template) | 1 μL | 2 μL | - |
| ddH2O | To 20 μL | To 50 μL | - |
Note: Keep template below 5% of the total PCR volume; 1 μL supernatant is recommended for a 20 μL reaction. A final primer concentration of 0.2-0.25 μM is recommended and may be optimized within 0.1-0.5 μM.
3. PCR cycling program
| Step | Temperature | Time | Cycles |
|---|---|---|---|
| Initial denaturation | 94°C | 5 min | 1 |
| Denaturation | 94°C | 10 sec | 35 |
| Annealing | 50-65°C | 20 sec | 35 |
| Extension | 72°C | 1 min | 35 |
| Final extension | 72°C | 5 min | 1 |
Note: Set the annealing temperature according to primer Tm, typically 2-5°C below the theoretical Tm. For fragments within 1 kb, an extension time of 1 min is recommended. PCR products may be loaded directly onto an agarose gel.
Storage
Lysis Buffer: 4°C, 1 year.
Neutralization Buffer, Direct PCR Master Mix (2×, with Dye): -20°C, 1 year. Avoid repeated freeze-thaw cycles.
Attention
1. Freshly collected leaf tissue is recommended. For samples stored long-term, keep them at −80°C and avoid repeated freeze–thaw cycles to prevent genomic DNA degradation, which may adversely affect PCR amplification efficiency and result reliability.
2. Young leaf tissue is preferred. If mature leaves must be used, avoid sampling from the main vein region.
3. This product is for R&D use only, not for drug, household, or other uses.
4. For your safety and health, please wear a lab coat and disposable gloves to operate.
Components
| Cat. No. | Product List | Components | HY-K0539-50 T | Storage |
|---|---|---|---|---|
| HY-K0539-A | Plant Tissue Direct PCR Reagent A | Lysis Buffer | 1.25 mL × 2 | 4°C, 1 year. |
| HY-K0539-B | Plant Tissue Direct PCR Reagent B | Neutralization Buffer Direct PCR Master Mix (2×, with Dye) |
500 μL 500 μL |
-20°C, 1 year. Avoid repeated freeze-thaw cycles. |
Please enter the correct Lot Number
