Plant Tissue Direct PCR Kit

 

Product Name	Cat.No.	Spec.
Plant Tissue Direct PCR Kit	G3313-01	100 rxns×20 μL

 

 

The kit is designed for direct PCR of different types of plant tissues. Simply treated plant tissues can be directly used as templates for PCR, without genome extraction. The product is compatible with common plant samples and polysaccharide polyphenol plant samples.

The 2×Plant Direct PCR Master Mix provided by this kit is specially optimized for direct amplification of plant tissue samples, which greatly improves the amplification efficiency and anti-inhibitor effect. In addition, the reagent contains all components except templates and primers, and the addition of dyes, the amplification products can be directly electrophoresis, which greatly simplifies the operation process. The simplicity and strong compatibility of this product can be used in molecular biology experiments such as rapid identification of transgenic plants and plant genotyping.

 

 

2×Plant Direct PCR Master Mix should be shipped with wet ice, and stored at -20°C; Other reagents are shipped with wet ice, and stored at 2~8°C; valid for up to 12 months.

 

 

Component Number	Component	G3313-01
G3313-1	2×Plant Direct PCR Master Mix	1 mL
G3313-2	Lysis Buffer A	5 mL
G3313-3	Lysis Buffer B	4 mL
G3313-4	Direct PCR Enhancer	500 μL
G3313-5	Nuclease-free Water	5 mL
Manual		One copy

 

 

1. PCR after lysis of plant tissue

1) Put the 5 mm² leaves (fresh and tender leaves are recommended) or 5 mg seeds (for hard seeds, you can first grind them into powder with a grinder or mortar) into a Nuclease-free centrifuge tube. Add 50 μL Lysis Buffer A, and mash the plant sample with the pipette tip (for example, the solution of tobacco leaves turns green after mashing, and the seed samples ground into powder are stirred evenly with the tip of a pipette). Place at room temperature for 5 min.

2) Add 40 μL Lysis Buffer B, mix well, centrifuge and collect the precipitate to the bottom of the tube. Remove the supernatant, dilute it 5 times with Nuclease-free Water and then use it as a template.

3) Recommend PCR reaction system:

Component	20 μL rxn	Final Concentration
2×Plant Direct PCR Master Mix	10 μL	1×
Forward Primer (10 μM)a	0.8 μL	0.4 μM
Reverse Primer (10 μM)a	0.8 μL	0.4 μM
Templateb	Variable (≤4 μL)
Nuclease-free Water	Add to 20 μL

 

a. The final concentration of primers ranges from 0.2 μM to 1.0 μM, and the recommended concentration of primers is 0.4 μM. Too few primers may lead to low amplification yield or no amplification, and excessive primers may lead to non-specific amplification.

b. In general, a good result can be obtained when the supernatant diluted 5 times by 2 μL is added to the 20 μL system. The amount of template can be adjusted according to whether it is difficult to lysis the plant sample, and the addition amount is not more than 4 μL.

2. Direct PCR of plant leaves (recommended PCR reaction system):

Component	50 μL rxn	Final Concentration
2×Plant Direct PCR Master Mix	25 μL	1×
Direct PCR Enhancer	10 μL
Forward Primer (10 μM)b	2 μL	0.4 μM
Reverse Primer (10 μM)b	2 μL	0.4 μM
Templatec	1 mm2 plant leaves
Nuclease-free Water	Add to 50 μL

a. Direct PCR of plant leaves without lysisis only suitable for amplification of fragments below 1 kb, and fresh and tender plant leaves are recommended.

b. The final concentration of primers ranges from 0.2 μM to 1.0 μM, and the recommended concentration of primers is 0.4 μM. Too few primers may lead to low amplification yield or no amplification, and excessive primers may lead to non-specific amplification.

c. It is recommended that the plant leaves added in the 50 μL system should be about 1mm², and cut the blade to 1mm² size with a punch or scissors.

3. Recommended PCR conditions:

Step	Temperature	Time	cycles
Initial Denaturationa	98°C	2 min~10 min	1
Denaturation	98°C	10 sec	35
Annealing	50~65°C	30 sec
Extensionb	72°C	10~15 sec/kb
Final extension	72°C	5~10 min	1
Hold	4~16°C	Forever

a. The Initial Denaturation time of plant tissue after lysis can be shortened to 2 min. It is recommended that the Initial Denaturation time of plant leaves without lysis is 10 min.

 

 

1. Please read the Product Manual carefully before use.

2. Try to use fresh plant leaves, otherwise the amplification efficiency will be affected.

3. When amplifying the fragments below 1 kb, 1mm² leaves can be directly used as templates. When amplifying the fragment of 1~2 kb, it is necessary to lysis first and then take its supernatant as a template.

4. 2×Plant Direct PCR Master Mix avoid repeated freezing and thawing and try to use it within one month after thawing.

5. For the preparation and sub-packaging of the reaction solution, please use a new pipette tip to avoid cross contamination.

 

For Research Use Only!