Fluorescence detection kit for detecting prawn enterocytozoon hepatopenaei (EHP) in China

A fluorescence detection and kit technology, which is used in the determination/inspection of microorganisms, DNA/RNA fragments, recombinant DNA technology, etc., can solve the problems of heavy workload, long detection time, strong subjectivity, etc. The effect of fast, real-time response results

Inactive Publication Date: 2017-12-19
INSPECTION & QUARANTINE TECH CENT OF XIAMEN ENTRY EXIT INSPECTION & QUARANTINE BUREAU +1
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Problems solved by technology

[0005] The purpose of the present invention is to solve the problems of long detection time, heavy workload, and strong subjectivity in the monitoring of breeding diseases in the existing detection methods, and provide high sensitivity, s

Method used

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  • Fluorescence detection kit for detecting prawn enterocytozoon hepatopenaei (EHP) in China
  • Fluorescence detection kit for detecting prawn enterocytozoon hepatopenaei (EHP) in China
  • Fluorescence detection kit for detecting prawn enterocytozoon hepatopenaei (EHP) in China

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Embodiment Construction

[0045] The technical solution of the present invention will be further specifically described below through specific examples.

[0046] In the present invention, unless otherwise specified, the raw materials and equipment used can be purchased from the market or commonly used in this field. The methods in the following examples, unless otherwise specified, are conventional methods in the art.

[0047] The invention includes tissue lysate, constant temperature fluorescence method reaction solution, positive control substance and negative control substance, wherein the positive control substance is a plasmid with shrimp hepatospora gene, and the negative control substance is ultrapure water without nucleic acid.

[0048] The components of the constant temperature fluorescence method reaction solution are as follows:

[0049] Primer EHP-OF 0.2 μM, primer EHP-OB 0.2 μM, primer EHP-IF 1.6 μM, primer EHP-IB 1.6 μM, primer EHP-LF 0.8 μM, primer EHP-LB 0.8 μM, Tris-HCl 20 mM (pH8.4 ...

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Abstract

The invention discloses a fluorescence detection kit for detecting prawn enterocytozoon hepatopenaei (EHP) in China and relates to detection of prawn diseases. The fluorescence detection kit is provided with a tissue lysis solution, a constant-temperature fluorescence process reaction solution, a positive reference substance and a negative reference substance. Application of the fluorescence detection kit comprises the following steps: extraction of prawn tissue DNA; constant-temperature fluorescence amplification of prawn EHP; result judgment of the constant-temperature fluorescence detection of prawn EHP. According to the fluorescence detection kit performing EHP detection through the constant-temperature fluorescence process, a positive sample can show a high-linearity amplification curve after being amplified by a constant-temperature fluorescence amplification apparatus, and also has a higher amplification rate; the result of a sensitivity experiment indicates that the fluorescence detection kit has higher sensitivity and can be applied to a quantitative experiment. A constant-temperature fluorescence primer with strong specificity, high sensitivity and high amplification efficiency is designed and developed for the genome DNA of prawn EHP; compared with a common LAMP primer, a loop primer is added, the amplification efficiency is higher, and the detection is faster.

Description

technical field [0001] The invention relates to the detection of prawn diseases, in particular to a fluorescence detection kit used for the detection of Enterospora hepatica of prawns in my country. Background technique [0002] Microsporidia are eukaryotic, unicellular obligate intracellular parasites, which are widely parasitic in invertebrates and vertebrates, and are important pathogens of some insects, aquatic animals and mammals. Enterocytozoon hepatopenaei (EHP), a new species of microsporidia, was first found in the slow-growing Penaeus monodon cultured in Thailand, and later in Litopenaeus vannamei, White shrimp and Penaeus sinensis etc. have been reported. This type of microsporidia mainly infects the hepatopancreas of prawns and destroys the ability of the tissue to attract nutrients. Although it does not cause a large number of prawns to die like WSSV, YHV, AHPND, etc., the problem of slow growth of prawns has brought huge losses to the prawn farming industry. ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
Inventor 徐淑菲谢会孔繁德罗达圣陈信忠高美玲张璜
Owner INSPECTION & QUARANTINE TECH CENT OF XIAMEN ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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