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Nucleic acid molecular hybridization detection method for silkworm nosema disease

A technology of nucleic acid molecular hybridization and silkworm microparticles, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of low sensitivity, poor accuracy, and long time consumption, and achieve high content and good quality.

Inactive Publication Date: 2004-10-06
CHENGDU TIANCHUANG BIO TECH
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Problems solved by technology

These three methods all have the following disadvantages: time-consuming, low sensitivity, poor accuracy, and inability to achieve early detection
At the same time, the existence of multiple variants of pathogenic protosporidium also increases the difficulty of the traditional detection method of microspores and microspores, further reducing the accuracy of detection

Method used

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  • Nucleic acid molecular hybridization detection method for silkworm nosema disease
  • Nucleic acid molecular hybridization detection method for silkworm nosema disease
  • Nucleic acid molecular hybridization detection method for silkworm nosema disease

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Embodiment Construction

[0016] Concrete operation steps of the present invention are as follows:

[0017] Extraction of Microsporidia DNA from Bombyx mori→polymerase chain reaction (PCR)→PCR product cloning and identification of recombinant plasmids→recombinant plasmid sequence analysis→specific identification of N.B. spores by DIG-labeled probes→detection sensitivity of N.B. Identification → Application of DIG-labeled probes in sericulture production.

[0018] 1. Extraction of DNA from Nosporum silkworm: extract with proteinase K phenol chloroform extraction. The silkworm fed with microsporidia is crushed and purified, and the operation is carried out as follows:

[0019]

[0020]

[0021] Microsporidia genomic DNA extracted by this method ( figure 1 ) content of up to 7 to 18 micrograms.

[0022] 2. Polymerase Chain reaction (PCR for short): According to the GenBank Sequence database of the National Center for Biotechnology Information (NCBI), the nucleotide sequences of all reported micro...

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Abstract

The present invention discloses a nucleic acid molecular hybridization detection method of domestic silkworm nosema disease, including the following steps: using nucleic acid molecular hybridizatino technology to extract DNA of domestic silkworm microsporidian, utilizing PCR to clone the PCR product into recombinant vector plasmid, then using N.P.PCR product and DIG marker to make molecular probe, and adopting DNA Dot blot to detect the pathogenic spore of silwkorm nosame disease. The various microsporidian genomes extracted by said invention are good in quality, high in content, and the designed primers are specific for N.B spora, and said primer and template DNA specificity are highly matched, and its detection accuracy is greater than 98%.

Description

Technical field: [0001] The invention relates to a detection method for biological diseases, in particular to a detection method for insect diseases, and more particularly to a nucleic acid molecular hybridization detection method for silkworm microparticle disease. Background technique: [0002] Pebrine disease is a silkworm disease caused by the parasitism of Nosema bombycis (N.B.) and is a devastating chronic infectious disease of silkworms. It is seriously harmful to silkworm production and cannot be cured for a long time. The disease has a wide range of epidemics and is difficult to diagnose and treat. It has historically dealt a devastating blow to the sericulture production of major sericulture countries in the world such as Europe, Japan, and China. Countries such as France and Italy are devastated because of the outbreak of silkworm microparticle disease, so the prevention and treatment of microparticle disease is a worldwide problem. [0003] In recent years, the ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 白燕川秦启联赵力宾
Owner CHENGDU TIANCHUANG BIO TECH
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