LAMP Fluorescent Master Mix HYB314
This product contains reaction buffer, enzymes mix of Bst DNA polymerase, lyophilized protectant and fluorescent dye components. The reaction buffer contains Mg2+, dNTP and other necessary components for amplification, users can directly mix the reaction buffer, enzymes mix, fluorescent dye, primer, and then add the template. If necessary, users can add the lyophilized protectant to prepare lyophilizable system.
Components
|
Components |
96 T |
960 T |
9600 T |
|
Loop-mediated Amplification Buffer |
1.2 mL |
4 mL×3 |
12 mL×10 |
|
Enzymes Mix |
192 μL |
1.92 mL |
1.92 mL×10 |
|
Lyophilized protectant |
0.96 mL×2 |
9.6 mL |
9.6 mL×10 |
|
25× Dye |
192 μL |
0.96 mL×2 |
0.96 mL×20 |
Storage
Store at -25～-15℃, valid for 12 months.
Instruction
1. Thaw the reaction buffer to be used at room Vortex briefly or invert tubes several times to mix thoroughly, then centrifuge to collect the liquid to the bottom of the tube.
2. Prepare reaction mix:this reagent can be prepared in two reaction systems, liquid reaction mix and Freeze-drying system mix.
1) Preparation of liquid reaction system
|
Component |
Volume |
|
Loop-mediated Amplification Buffer |
10 uL |
|
25× Dye |
2 μL |
|
Enzymes Mix |
2 μL |
|
10×Primer Mix* |
5 μL |
|
Template DNA** |
X μL |
|
Nuclease-free Water |
Up to 50 μL |
2) Preparation of freeze-dried reaction system
① Preparation of freeze-dried system
|
Component |
Volume |
|
Loop-mediated Amplification Buffer |
12.5 uL |
|
Lyophilized protectant |
10 μL |
|
25× Dye |
2 μL |
|
Enzymes Mix |
2 μL |
|
Nuclease-free Water |
Up to 50 μL |
② Freeze-drying
The prepared Mix was freeze-dried in a 50μL system.
③Prepare reaction mix
|
Component |
Volume |
|
Freeze-dried product |
1 piece |
|
10×Primer Mix* |
5 μL |
|
Template DNA** |
x μL |
|
Nuclease-free Water |
Up to 50 μL |
*10× Primer Mix concentration: 16 μM FIP/BIP, 2 μM F3/B3, 4 μM Loop F/B;
**It is recommended to dissolve the nucleic acid template in DEPC water.
3. Reaction program
Set the parameters on the PCR instrument (such as ABI7500) as follows:
|
Temperature |
Time |
Cycles |
|
65℃ |
60 s **Collect fluorescent signal |
30 |
Notes
1. Salt may appear in the bottom of the buffertube, vortex briefly or invert tubes several times to mix thoroughly at room temperature.
2. The reaction temperature can be optimizedbetween 62°C and 68°C according to the primer
3. The experiment shall be conducted in astandardized manner, including the preparation of reaction system, sample treatment and sample
4. It is suggested to prepare reaction system inthe ultra-clean table and add templates in the fume hood of other rooms to avoid false.