LAMP Turbo绿色预混液

LAMP Turbo绿色预混液，Saphir LAMP Turbo GreenMaster，用于快速等温DNA扩增并含绿色荧光DNA染料的预混液。更多视频请关注视频号【艾维缔】。哔哩哔哩【IVDSHOW】。抖音【军哥聊表观】。

关键词

LAMP Turbo绿色预混液

所属分类

分子生物学

目录号

PCR393

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For general laboratory use.

Please centrifuge briefly before opening (volume ≤2 ml).

Shipping: shipped on gel packs

Storage Conditions: store at -20 °C
store dark
Short term storage (up to 3 months) at 4 °C possible.

Shelf Life: 12 months

Form: liquid

Concentration: 2x conc.

Spectroscopic Properties: λ_exc 495 nm, λ_em 520 nm (dye bound to DNA)

Description:
Saphir LAMP Turbo GreenMaster is a complete 2x conc. master mix for isothermal amplification of DNA. The mix is based on a genetically enhanced Bst polymerase of the next generation. The mixe is the ideal choice for ultra-fast and robust amplification of DNA at constant temperature (60 to 65 °C). The enzyme shows high strand displacement activity and generates an ampliﬁcation factor of up to 10⁹ which is comparable to approx. 30 cycles in a PCR assay. The polymerase is 2-3x faster compared to Saphir Bst Polymerase (#PCR-389/#PCR-387) and allows detection of a target gene within 5-10 minutes.

Content:
Saphir LAMP Turbo GreenMaster
Saphier Bst Turbo Polymerase, dNTPs, reaction buffer, glycerol, green-fluorescent DNA stain, additives and stabilizers

PCR-grade water

Detection
The mix contains a green-fluorescent DNA stain that intercalates into DNA during the amplification process and allows the direct quantification of target DNA by fluorescence detection (analogous to real-time PCR).
The mix can be combined with ROX reference dye (#PCR-351) to allow a signal normalization in real-time PCR instruments that are compatible with the evaluation of the ROX signal.

Assay design
Isothermal ampliﬁcation is an extremely sensitive detection method and care should be taken to avoid contamination of set-up areas and equipment with DNA of previous reactions. A problem may be ampliﬁcation in no-template controls due to carry-over contamination or ampliﬁcation of unspecifically annealed primers or primer dimer formations.

Primer design
Typically, 4 different primers are used to identify 6 distinct DNA regions allowing the specific amplification of a target gene. An additional pair of primers further accelerates the amplification allowing to cut down the total detection time to 5-10 min.
The manual design of primers may be challenging due to the complex reaction sequence. To simplify the design process the use of a primer design software is recommended.
As sensitivity and non-template ampliﬁcation of in-silico designed primers may vary, the evaluation of 2 - 4 real primer sets before choosing a ﬁnal set is recommended.

Assay set-up
A reaction volume of 20-50 μl is recommended for most applications. Pipet with sterile ﬁlter tips and perform the set-up in an area separate from DNA preparation or analysis. No-template controls should be included in all ampliﬁcations.
First, prepare a 10x conc. primer pre-mix. Second, set-up the isothermal ampliﬁcation assay:

component	stock conc.	final conc.	20 μl	50 μl
Saphir LAMP Turbo GreenMaster	2x	1x	10 μl	25 μl
Primer Mix	10x	1x	2 μl	5 μl
Template DNA		<500 ng/assay	x μl	x μl
PCR-grade Water			fill up to 20 μl	fill up to 50 μl

Use a specific detection instrument for isothermal amplification or a real-time PCR cycler to run the assays
Set the instrument to a constant incubation temperature between 60 to 65°C (depending on the primer annealing temperature)
Measure the fluorescence intensity at an interval of 1 min for up to 30 min.

Trouble shooting
If amplification in no-template controls occurs the following points should be reviewed.

Cross contamination from environments

Clean equipment and areas with “DNA Away” solution
Replace reagent stocks and pre-mixes with new components
Stop reactions at an earlier point of time before non-template amplification occur

Carry-over contamination from previous reaction products

Avoid opening reaction vessels after amplification
Use separate preparation area and equipment if post-reaction processing is necessary

Non-template amplification from primers

Increase incubation temperature stepwise by 1-2 °C
Design a new set of primers for the target sequence

产品组分

组分内容	型号	规格	储存温度
LAMP Turbo绿色预混液	PCR393-01	2x1.25ml	-20°C
LAMP Turbo绿色预混液	PCR393-02	10x1.25ml	-20°C
操作手册	1	1	室温

*注意：在使用之前，应将溶液离心至管底。

文件资源

营销中心

注意事项

保存建议	推荐蓝冰或冰袋运输。当您收到产品后，按照说明书建议保存各组分。
警告	本品仅供科研使用，请勿用于临床与诊断。

FAQ
Evaluating Commercial Loop-Mediated Isothermal Amplification Master Mixes for Enhanced Detection of Foodborne Pathogens

Ana Costa-Ribeiro, Alexandre Lamas, ..., Katrina Counihan

Foods | 2024 May 24