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Development of a Rapid Detection Method for Potato virus X by Reverse Transcription Loop-Mediated Isothermal Amplification.

Jeong J, Cho SY, Lee WH, Lee KJ, Ju HJ - Plant Pathol. J. (2015)

Bottom Line: RT-LAMP for the detection of Potato virus X (PVX) was developed and compared with conventional reverse transcription polymerase chain reaction (RT-PCR) to demonstrate its advantages over RT-PCR.Based on real-time monitoring, RT-LAMP detected PVX around 30 min, compared to 120 min for RT-PCR.By adding a fluorescent reagent during the reaction, the extra step of visualization by gel electrophoresis was not necessary.

View Article: PubMed Central - PubMed

Affiliation: Department of Agricultural Biology, College of Agriculture & Life Sciences, Chonbuk National University, Jeonju-si 561-756, Korea.

ABSTRACT
The primary step for efficient control of viral diseases is the development of simple, rapid, and sensitive virus detection. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) has been used to detect viral RNA molecules because of its simplicity and high sensitivity for a number of viruses. RT-LAMP for the detection of Potato virus X (PVX) was developed and compared with conventional reverse transcription polymerase chain reaction (RT-PCR) to demonstrate its advantages over RT-PCR. RT-LAMP reactions were conducted with or without a set of loop primers since one out of six primers showed PVX specificity. Based on real-time monitoring, RT-LAMP detected PVX around 30 min, compared to 120 min for RT-PCR. By adding a fluorescent reagent during the reaction, the extra step of visualization by gel electrophoresis was not necessary. RT-LAMP was conducted using simple inexpensive instruments and a regular incubator to evaluate whether RNA could be amplified at a constant temperature instead of using an expensive thermal cycler. This study shows the potential of RT-LAMP for the diagnosis of viral diseases and PVX epidemiology because of its simplicity and rapidness compared to RT-PCR.

No MeSH data available.


Related in: MedlinePlus

Effect of loop primers on RT-LAMP amplification of the PVX CP gene. Real-time measurement of optical density with a real-time turbidimeter was used to monitor RT-LAMP amplification with or without loop primers.
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f4-ppj-31-219: Effect of loop primers on RT-LAMP amplification of the PVX CP gene. Real-time measurement of optical density with a real-time turbidimeter was used to monitor RT-LAMP amplification with or without loop primers.

Mentions: RT-LAMP reactions were performed using primer set A with or without loop primers to examine the effect of loop primers on shortening the reaction time according to the turbidity of reaction mixtures with or without loop primers. The results showed that the time required for the initiation of RT-LAMP amplification was 13 or 26 min with or without loop primers, respectively (Fig. 4). Evidently, the use of loop primers accelerated the amplification of PVX, reducing the reaction time to almost half.


Development of a Rapid Detection Method for Potato virus X by Reverse Transcription Loop-Mediated Isothermal Amplification.

Jeong J, Cho SY, Lee WH, Lee KJ, Ju HJ - Plant Pathol. J. (2015)

Effect of loop primers on RT-LAMP amplification of the PVX CP gene. Real-time measurement of optical density with a real-time turbidimeter was used to monitor RT-LAMP amplification with or without loop primers.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4564147&req=5

f4-ppj-31-219: Effect of loop primers on RT-LAMP amplification of the PVX CP gene. Real-time measurement of optical density with a real-time turbidimeter was used to monitor RT-LAMP amplification with or without loop primers.
Mentions: RT-LAMP reactions were performed using primer set A with or without loop primers to examine the effect of loop primers on shortening the reaction time according to the turbidity of reaction mixtures with or without loop primers. The results showed that the time required for the initiation of RT-LAMP amplification was 13 or 26 min with or without loop primers, respectively (Fig. 4). Evidently, the use of loop primers accelerated the amplification of PVX, reducing the reaction time to almost half.

Bottom Line: RT-LAMP for the detection of Potato virus X (PVX) was developed and compared with conventional reverse transcription polymerase chain reaction (RT-PCR) to demonstrate its advantages over RT-PCR.Based on real-time monitoring, RT-LAMP detected PVX around 30 min, compared to 120 min for RT-PCR.By adding a fluorescent reagent during the reaction, the extra step of visualization by gel electrophoresis was not necessary.

View Article: PubMed Central - PubMed

Affiliation: Department of Agricultural Biology, College of Agriculture & Life Sciences, Chonbuk National University, Jeonju-si 561-756, Korea.

ABSTRACT
The primary step for efficient control of viral diseases is the development of simple, rapid, and sensitive virus detection. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) has been used to detect viral RNA molecules because of its simplicity and high sensitivity for a number of viruses. RT-LAMP for the detection of Potato virus X (PVX) was developed and compared with conventional reverse transcription polymerase chain reaction (RT-PCR) to demonstrate its advantages over RT-PCR. RT-LAMP reactions were conducted with or without a set of loop primers since one out of six primers showed PVX specificity. Based on real-time monitoring, RT-LAMP detected PVX around 30 min, compared to 120 min for RT-PCR. By adding a fluorescent reagent during the reaction, the extra step of visualization by gel electrophoresis was not necessary. RT-LAMP was conducted using simple inexpensive instruments and a regular incubator to evaluate whether RNA could be amplified at a constant temperature instead of using an expensive thermal cycler. This study shows the potential of RT-LAMP for the diagnosis of viral diseases and PVX epidemiology because of its simplicity and rapidness compared to RT-PCR.

No MeSH data available.


Related in: MedlinePlus