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Hieff Unicon® Universal TaqMan multiplex qPCR master mix

Product SKU

Specifications

Price

11211ES03

1 mL

$221.10

11211ES08

5 mL

$885.02

11211ES60

100 mL

Product Description

This product is a 2×reagent, which can achieve up to four times qPCR reaction. The product contains the antibody bolck Taq enzyme to improves the sensitivity and specificity during amplification. At the same time, optimized reaction buffer can improve the amplification efficiency and promote the effective amplification of low concentration template. This product can be used for genotyping and multiple quantitative analysis of genes.

Package Information

Contents No.

Name

11211ES03 (1 mL)

11211ES08 (5 mL)

11211ES60 (100 mL)

11211-A

2×Hieff Unicon® Universal TaqMan multiplex qPCR master mix

1 mL

1 mL×5

1 mL×100

Shipping and Storage

Transport in ice packs,it can be stored at -20℃ for 1 years.

Reaction composition

Component

Volume (μL)

Final Concentration

2×Hieff Unicon® Universal TaqMan multiplex qPCR master mix

12.5

Primer mix (10 μM)

X

0.1 μM-0.5 μM

Probe mix (10 μM)

X

50 nM-250 nM

Rox reference dye

0.4

Template DNA/cDNA

1-10

-

ddH2O

up to 25

-

Note: Be sure to mix well before use, avoid excessive bubbles caused by violent vibration.

a) Primer concentration: Primer Mix including multiplex primer, each primer’s finally concentration is 0.2 μM usually, depending on the situation optimal primer concentration may be between 0.1 and 0.5 μM.

b) Probe concentration: Probe Mix including multiplex probe labeling difference Fluorescent group, depending on the situation optimal probe concentration may be between 0.1 and 0.5 μM.

c) Rox reference dye: It is used in real time PCR instrument such as Applied Biosystems to correct the error of fluorescence signal generated between pores; this product does not contain Rox reference dye. If necessary, the product Cat No.10200 is recommended.

d) Template dilution: qPCR is highly sensitive and it is recommended to dilute the template. If the template is the original solution of cDNA, the volume of the template does not exceed 1/10 of the total volume.

e) Reaction composition: 25 μL, 30 μL or 50 μL are recommended to ensure the effectiveness and repeatability of target gene amplification.

f) System preparation: Please prepare in the ultra-clean working table, and pipettor and reaction tube without nuclease residue; it is recommended to use the gun head with filter element. Avoid cross contamination and aerosol contamination.

Thermal cycling protocol

Stage

Temperature

Time

Cycles

Pre-denaturation

95℃

5 min

1

Denaturation

95℃

15 sec

45

Annealing/Extension

60℃

30 secb

45

Note

a) Annealing/Extension: The temperature and time can be adjusted according to the experimental requirements.

b) Fluorescence signal acquisition: Please set the experimental procedure according to the requirements of the instrument manual. The time settings of several common instruments are as follows:

20 sec: Applied Biosystems 7700, 7900HT, 7500 Fast

31 sec: Applied Biosystems 7300

32 sec: Applied Biosystems 7500

Important points before starting

1. About operation

1) Please make sure that the product is completely thawed and well mixed before use. After being centrifuged and collected to the bottom of the tube, put it on ice for standby.

2) In order to avoid cross contamination between samples and aerosol pollution, it is recommended to prepare the reaction system in the ultra-clean platform.

3) Avoid repeated freezing and thawing during use.

4) For your safety and health, please wear lab clothes and disposable gloves.

2、Experimental methods

1) Primer design

a) Primers must be specific.

b) The Tm value of primer must between 58 and 60℃, and the Tm difference of each primer should be controlled within 1-2℃.

c) The length of primers is generally controlled between 15-30 bp.

2) Probe design

a) The Tm value of TaqMan probe was about 10℃ (68-70℃) higher than that of corresponding primers.

b) There was no dimer between TaqMan probe and primer.

15221439726

15221439726