LAMP绿色冻干预混液

LAMP绿色冻干预混液，Saphir LAMP GreenMaster Lyophilisate，含绿色荧光DNA染料的快速等温DNA扩增冻干粉。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绿色冻干预混液

所属分类

分子生物学

目录号

PCR398

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For general laboratory use.

Shipping: shipped at ambient temperature

Storage Conditions: store at ambient temperature
Store in an aluminium-coated bag or in a dry place.
Lyophilisates may hydrate at humidity levels >70 % when sealing is opened.

Shelf Life: 6 months

Description:
Saphir LAMP GreenMaster Lyophilisate is a freeze-dried master mix for isothermal amplification of DNA using LAMP (Loop-mediated Isothermal Amplification). The mix is based on the activity of Bst polymerase and is the ideal choice for 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.
Saphir LAMP GreenMaster Lyophilisate allows detection of a target gene within 10-20 minutes.

Content:
Saphir LAMP GreenMaster Lyophilisate
Saphir Bst Polymerase, dNTPs, reaction buffer, Green DNA intercalator dye, additives and stabilizers in freeze-dried format for 20 μl final assay volume

PCR grade water

Handling
The lyophilisates are provided in low-profile (0.1 ml) 96-well plates with optically clear caps, whereby each well contains reaction mix for a final volume of 20 μl. The plates can be easily divided into 8-well strips and further segmented by cutting, allowing compatibility with a variety of PCR cyclers.
The lyophilisate combines highest performance with convenience of use and stability. There is no need for freezing, thawing or pipetting on ice. The few remaining pipetting steps minimize the risk of errors or contaminations.
Each vial contains all components (except primers and template) required for a 20 μl LAMP assay.
To perform the assay, only fill up the vials with a primer mix and add DNA template.
The lyophilisate can also be used with ROX reference dye in PCR instruments that are compatible with the evaluation of the ROX signal. In this case, the ROX dye (#PCR-351) should be added as 1x concentration to the PCR reaction.

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 10-20 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
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.	Volumne 1x 20 μl Assay
Primer Mix	10x	1x	2 μl
Template DNA		<500 ng/assay	x μl
PCR-grade Water			fill up to 20 μl

Dispensing the master mix
Vortex the primer/probe mix thoroughly to assure homogeneity.
Dispense 20 μl to each PCR tube or well of the plate.

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绿色冻干预混液	PCR398-01	192次	-20°C
LAMP绿色冻干预混液	PCR398-02	960次	-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。

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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。