Detection of Plant Nematodes by Loop-Mediated Isothermal Amplification

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Despite its good specificity and sensitivity, the requirement of high precision of thermal cycling in PCR prevents this powerful method for being widely used as a routine diagnostic tool. The recent invention of loop-mediated isothermal amplification (LAMP) provides a new alternative for molecular diagnosis, which requires the design of four to six primers, including external primers and internal primers. It is only necessary to use Bst DNA polymerase, with strand displacement and recognition elongation effect, to amplify the target fragment in large and rapid quantities at 60 - 65°C.

Powered by our professional scientists and their years of field experience, Lifeasible provides detection of plant nematodes by LAMP and crossing priming amplification (CPR) to assist our customers in the identification of plant nematodes. With our cutting-edge platforms, we can achieve satisfactory results for our clients worldwide.

Detection of Plant Nematodes by LAMP

Since its development, LAMP technology has been widely used and achieved good results in many aspects of life science. In recent years, LAMP technology has been gradually applied to the field of plant pathogen detection, including nematodes.
In LAMP amplification products, SYBR Green I dye is added to facilitate the visual display of detection results.

Fig.1 Visual inspection and lateral-flow strips used for the detection of LAMP products.

Lifeasible provides detection of plant nematodes with LAMP, which is simple to operate, low cost, and low equipment demand, and can be used for field detection with fluorescent samples without expensive instruments and equipment.

Workflows of LAMP Detection

Steps	Operation Methods
DNA Extraction	Plants' nematodes are isolated from culture by the Baermann funnel technique and washed three times with phosphate-buffered saline-Twain. After incubation with 100 µl of extraction buffer at 55°C for 20 min, the lysate is extracted once with PCI (phenol / chloroform / isoamyl alcohol), followed by ethanol precipitation, and finally dissolved in 50 µl Tris-EDTA buffer.
Primer Design	The specific primers for the LAMP reaction are designed using the software program.
LAMP Reaction	The reaction is performed in the reaction mixture containing extracted DNA solution, primers, reaction mix, Bst DNA polymerase, and fluorescent detection reagent. The reaction mixture is incubated at 63°C for 60-120 min and terminated by incubation at 80°C for 2 min. LAMP amplicons are detected by color changes of the reaction solution under UV light.
Lateral-Flow Strip Detection	After the LAMP reaction, 10 µl of FITC-labeled probe designed to hybridize to an internal region of the target sequence is added to the reaction mixture and incubated at 95°C for 5 min. The reaction mixture is diluted with 100 µl of running buffer and applied directly to detect strips.
Sensitivity Test	A plasmid DNA containing the target sequence is made using the pGEM-T Easy System.

Detection of Plant Nematodes by CPR

Based on the LAMP technology, the CPR appears to detect plant nematodes.
We provide detection of plant nematodes with CPR, which has the advantages of high sensitivity, strong specificity, easy to interpretation, simple detection equipment, low one-time investment, easy to popularize, and so on.

Lifeasible offers the detection services of plant nematodes for the convenience of your research. Our goal is to help our clients achieve meaningful results through powerful and consistent approaches. If you are interested in our services or have any questions, please feel free to contact us or make an online inquiry.

Our products/services are For Research Use Only. Not For Clinical Use!

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