Indian Journal of Agricultural Research

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Research Article

Indian Journal of Agricultural Research, volume 53 issue 4 (august 2019) : 447-452

Loop Mediated Isothermal Amplification (LAMP) for Nosema bombycis diagnosis by Small subunit Ribosomal RNA (SSU rRNA) gene

S. Kampliw, M. Monthatong
1Department of Biology, Faculty of Science, Khon Kaen University, Khon Kaen, 40002 Thailand.
Cite article:- Kampliw S., Monthatong M. (2019). Loop Mediated Isothermal Amplification (LAMP) for Nosema bombycis diagnosis by Small subunit Ribosomal RNA (SSU rRNA) gene. Indian Journal of Agricultural Research. 53(4): 447-452. doi: 10.18805/IJARe.A-430.

ABSTRACT

In present study, Loop Mediated Isothermal Amplification (LAMP) assay was conducted for diagnosis Nosema bombycis, the pebrine disease pathogen in domestic silkworm, Bombyx mori. Nine isolates of N. bombycis were collected from infected silkworms in rearing areas in Khon Kaen province, Thailand. N. bombycis genomic DNAs were extracted by boiling method and used as templates in LAMP and PCR reactions. A LAMP primer set was designed specific to 
N. bombycis small subunit ribosomal RNA (SSU rRNA) gene. The results revealed that the optimal condition was constantly performed at 63oC for 1 hour. The product was directly visualized by naked eye and confirmed with agarose gel eletrophoresis. LAMP assay is more sensitive than traditional PCR, since LAMP was able to detect the least 10 spores/ml while PCR needs 100 spores/ml. In addition, the present novel LAMP primer set was specific only to N. bombycis proven by the negative results when other B. mori pathogen DNAs were tested. In conclusion, the LAMP assay demonstrated a great potential alternative method in diagnosis N. bombycis with high sensitivity, rapidity and accuracy which can apply for pebrine disease surveillance. 

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