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Method for identifying physiological sex of fish at early development stage by using 5SrRNA

A developmental stage, fish technology, applied in the field of fish biological sex molecular identification, can solve the problems of waste of resources, increase costs, hinder the growth of industrial scale, etc., and achieve the effects of low cost, high accuracy and simple operation.

Pending Publication Date: 2021-10-22
HUAZHONG AGRI UNIV
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Problems solved by technology

This is unacceptable for large or economically valuable fish
More importantly, breeding too many male individuals is a huge waste of resources for industries such as sturgeon, which not only increases costs, but also hinders the scale growth of industries
In addition, the sex of fish is easily affected by environmental stress conditions such as stress temperature, high density, low dissolved oxygen, etc., so that the genetic sex and phenotypic sex are inconsistent, that is, there are XX male fish or XY female fish Individuals, while sex-specific molecular markers can only determine the genetic sex of the individual, but not the biological sex

Method used

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  • Method for identifying physiological sex of fish at early development stage by using 5SrRNA
  • Method for identifying physiological sex of fish at early development stage by using 5SrRNA
  • Method for identifying physiological sex of fish at early development stage by using 5SrRNA

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Embodiment 1

[0028] Combine below figure 1 with figure 2 The present invention is described in further detail:

[0029] A kind of method utilizing 5S rRNA to identify the physiological sex of fish early stage of development, its steps are:

[0030] (1) Acquisition of gonad tissue samples. The gonad tissues of yellow catfish just after sex differentiation, bluegill sunfish, golden perch, and largemouth bass at the gonadal development stage were collected and stored in RNAlater preservation solution (Sigma-Aldrich) immediately, and placed in a refrigerator at 4°C for about 12 hours before transferring to Store in -80°C refrigerator.

[0031] (2) Extraction of total RNA from gonad tissue. After homogenizing 20 mg of gonad tissue, use animal tissue total RNA rapid extraction kit (Kangwei Century or Thermofisher Ambion, etc.) to extract total RNA from gonad tissue, and adjust the RNA concentration to 50 ng / μl.

[0032] (3) Total RNA quality identification. In the agarose gel imaging of t...

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Abstract

The invention discloses a method for identifying the physiological sex of fish at an early development stage by using 5S rRNA, which comprises the following steps: (1) extracting the total RNA of fish gonad tissues by using a method for conventionally extracting the total RNA of animal cells by using a total RNA extraction kit or a TRIzol reagent, determining an OD260 / 280 value by using a nucleic acid analyzer, and judging the RNA purity, and then separating total RNA by using an agarose gel electrophoresis method, and observing the brightness and distribution of 5S rRNA, 18S rRNA and 28S rRNA; (2) carrying out microcapillary electrophoresis on the extracted gonad total RNA, and calculating the proportion of 5S rRNA in the total RNA according to the RNA type coverage area obtained by nucleic acid analysis software; (3) according to the proportion of the 5S rRNA in the total RNA, judging the physiological male and female of the fish at the early development stage, when the proportion of the 5S rRNA is smaller than or equal to 9.0%, judging the fish as a testis tissue, and when the proportion of the 5S rRNA is larger than or equal to 12%, judging the fish as an ovarian tissue. The method is easy to implement and simple and convenient to operate, the physiological sex identification of more than 100 fish individuals is realized within 12 hours, and the physiological sex of the fish is determined.

Description

technical field [0001] The invention belongs to the technical field of molecular identification of fish biological sex, and more specifically relates to a method for identifying the biological sex of fish in the early development stage by using 5S rRNA. Background technique [0002] Physiological sex identification at the early developmental stage of fish is of great value for research fields related to gonad development, fish resource investigation, and aquaculture. At the same time, for large and medium-sized fish (such as sturgeon, huso, and salmon), the identification of physiological sex in the early stage can avoid subsequent continuous investment in unnecessary sex (such as sturgeon males), so as to carry out targeted research. It is of great significance to improve the economic efficiency of farming by reducing the cost of farming on the ground. [0003] However, so far, the identification of physiological sex in the early developmental stages of fish completely dep...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6888C12Q1/6879
CPCC12Q1/6888C12Q1/6879C12Q2600/158C12Q2600/178
Inventor 沈志刚别汉晴胡晨昊李波于跃鲁子怡耿龙慧樊启学袁勇超
Owner HUAZHONG AGRI UNIV
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