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Method for separating and purifying myxobolus honghuensis polar capsule

A technology for separation, purification and iodine algae, which is applied in the biological field, can solve the problems of lack of technical specifications, high cost, complicated operation and the like, and achieves the effects of convenient operation, low cost and simple process

Active Publication Date: 2019-07-26
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The existing polar capsule extraction method is realized by microfluidic chip technology based on quadrupole electrode array, but this method is complicated to operate, high in cost, and lacks technical specifications

Method used

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  • Method for separating and purifying myxobolus honghuensis polar capsule
  • Method for separating and purifying myxobolus honghuensis polar capsule
  • Method for separating and purifying myxobolus honghuensis polar capsule

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Embodiment 1

[0023] The present invention is a separation method based on gradient centrifugation, and its key elements are centrifugal medium (sucrose, Percoll, etc.) and gradient composition. In order to determine the optimal method for the separation and purification of Honghu iodine polar capsules, a series of explorations and optimizations were carried out on the key elements of the gradient centrifugation steps:

[0024] Scheme 1 used sucrose with low cost as the centrifugation medium, and carried out the first exploration of polar capsule separation. The steps are as follows:

[0025] A. Take 5 mL of Honghu iodine spore fragmentation solution and add it to the bottom of a 50 mL centrifuge tube;

[0026] B. Through the syringe, inject the sucrose solutions with mass percentages of 40%, 45%, 50%, 55%, and 60% into the centrifuge tube in sequence, each layer is 8mL;

[0027] C, 3500rpm speed, centrifugal 15min;

[0028] Results: After centrifugation, the ratio of polar capsules to sh...

Embodiment 2

[0043] The separation and purification method of Honghu iodine polar capsules includes tissue sample homogenization, spore purification, ultrasonic crushing, Percoll gradient centrifugation, polar capsule recovery, secondary purification, resuspension and cleaning of recovered liquid, and the specific steps are as follows:

[0044] A. Tissue sample homogenization: cut out the crucian crucian throat that was found to contain Honghu iodine cysts by microscopic examination, put it into a hand-held homogenizer, and then add 0.1M PBS buffer to grind thoroughly until there is no obvious tissue in the homogenizer. organization block;

[0045]B. Purification of spores: filter the homogenate with double layers of medical gauze, collect the filtrate; centrifuge at 2500rpm for 15min, wash twice with 0.1M PBS buffer, resuspend the spores to 8mL with PBS, and add to the bottom of a 50mL centrifuge tube ; The sucrose solutions with mass percentages of 30%, 40%, 50%, and 60% were injected in...

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Abstract

The invention discloses a method for separating and purifying a myxobolus honghuensis polar capsule. The method includes tissue sample homogenization, spore purification, ultrasonic pulverization, Percoll density gradient centrifugation, polar capsule recovery, secondary purification and recovery liquid resuspension cleaning. According to the method, the difference in mass specific gravity of thepolar capsule and other impurities is utilized to achieve the separation of the polar capsule and the impurities. 50%, 70% and 90% Percoll separation media are utilized to perform density gradient centrifugation on a spore breaking liquid, and the complete polar capsule with the purity close to 100% can be obtained between the 50% and 70% Percoll solution layers and in the middle of the 70% Percoll separation medium. The method provides a new way for the source of the myxobolus honghuensis polar capsule, and the purified polar capsule can help to study the biological characteristics of the myxobolus honghuensis, analyze the weak links of life history, clarify the pathogenic mechanism and establish the early detection means and the like.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a method for separating and purifying polar capsules of Honghu iodine hyacinthia. Background technique [0002] Myxozoans are common parasites in farmed fish, and some species can cause major diseases of farmed fish. Among them, Honghu iodine worm is the pathogen of heterogeneous gibel crucian carp's "throat sporosis". It will form bean curd residue-like cysts in the throat of crucian carp, deprive the host of nutrients, hinder the fish from eating and cause mechanical compression, eventually leading to the death of the fish. The mortality rate of "throat spore disease" can exceed 90%, which seriously endangers the development of heterogeneous gibel crucian carp aquaculture industry. At present, the prevention and control of "laryngosporia" is facing several bottlenecks. On the one hand, the biological characteristics and pathogenic mechanism of Honghu iodophora...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/10C12N1/02C12R1/90
CPCC12N1/10C12N1/02
Inventor 顾泽茂郭庆祥翟艳花柳阳
Owner HUAZHONG AGRI UNIV
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