Efficient induction method and microsatellite molecular marker identification method of Nile tilapia pseudo-female fishes

A Nile tilapia and molecular marker technology, which is applied in the field of efficient induction of Nile tilapia pseudo-female and identification of microsatellite molecular markers, can solve the problems of indistinguishability, long breeding cycle, time-consuming and labor-intensive, etc.

Inactive Publication Date: 2017-02-15
SUN YAT SEN UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, since the sex chromosome composition of fish is in a primitive state, it is morphologically difficult to distinguish between Y and Z, X chromosomes
Due to the lack of means to accurately determine the genetic sex of tilapia, the use of traditional selective breeding to obtain all-male fish has the problems of long breeding cycle, time-consuming and labor-intensive problems

Method used

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  • Efficient induction method and microsatellite molecular marker identification method of Nile tilapia pseudo-female fishes
  • Efficient induction method and microsatellite molecular marker identification method of Nile tilapia pseudo-female fishes
  • Efficient induction method and microsatellite molecular marker identification method of Nile tilapia pseudo-female fishes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Embodiment 1: the preparation method of estrogen diethylstilbestrol (DES) opening feed

[0032] Firstly, the estrogen diethylstilbestrol (DES) was dissolved in 500 mL of absolute ethanol according to the required amount (0.5 g or 1 g) of the experiment, and then the absolute ethanol containing DES was fully stirred with 1 kg of feed for small fish. Refrigerate overnight at 4 degrees to allow small fish to fully absorb DES. Put the small fish opening feed added with DES into the fume hood to allow the absolute ethanol to fully volatilize. Dried to prepare 0.5g / kg, 1g / kg DES-containing open feed, then sealed with a plastic bag, and stored at low temperature for subsequent use.

Embodiment 2

[0033] Example 2: Family Construction and Estrogen Treatment

[0034] In June 2015, a pair of high-quality Nile tilapia broodstock (genetic male parent and genetic female parent) were selected at Guangzhou Wulonggang Aquatic Development Co., Ltd. to successfully construct a full-sib family (1356 in total). The experimental fish were bred and treated in Guangzhou Wulonggang Aquatic Products Development Co., Ltd.

[0035] The family was randomly divided into 3 groups (2 treatment groups and 1 control group) regardless of gender for breeding management (Table 1). Two of the treatment groups were fed the open feed containing DES when they started eating after hatching, and were fed continuously for 3 weeks to promote the development of genetic males into pseudo females. The control group was fed with small fish opening feed without adding estrogen.

[0036] Table 1 Nile tilapia full-sib family DES treatment phenotype sex statistics

[0037]

[0038] After the Nile tilapia gr...

Embodiment 3

[0039] Example 3: Genomic DNA source and preparation and marker screening

[0040] 1. Genomic DNA sample extraction from parents and offspring

[0041] The samples used in this experiment for the identification of sex-linked molecular markers of pseudo-female fish include: female and male parents in Example 2, 96 female (phenotype) individuals and 96 male (phenotype) individuals in the control group, and 1 treatment group 96 female (phenotype) individuals.

[0042] 2. DNA extraction method

[0043] (1) Parental DNA

[0044] DNA was extracted using Dongsheng Genomic DNA Rapid Extraction Kit, and the extracted DNA was diluted 5 times and stored at -20°C for later use.

[0045] (2) offspring DNA

[0046] a. Take a 1.5mL 96-well plate, put it on ice, add 300μL of STE buffer (10mM Tris pH8.0, 50mM EDTA pH 8.0, 200mM NaCl, 0.5% SDS), 50μL proteinase K (20mg / mL) to each well ;

[0047] b. Cut 3*3mm with sterilized scissors 2 Fish fin samples of the same size, placed on filter ...

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Abstract

The invention discloses an efficient induction method and a microsatellite molecular marker identification method of Nile tilapia pseudo-female fishes. Different doses of estrogen diethylstilbestrol (DES) are used for sex reversal processing of different groups of Nile tilapia fishes to establish the efficient induction method of the Nile tilapia pseudo-female fishes; and a newly-developed microsatellite molecular marker tlp-23-sd and a primer thereof are further used for identification of the Nile tilapia pseudo-female fishes to establish the microsatellite molecular marker rapid identification method of the pseudo-female fishes, wherein the microsatellite molecular marker tlp-23-sd is closely linked with the sex. The efficient induction method is high in pseudo-female fish induction efficiency, and the microsatellite molecular marker rapid identification method is rapid, accurate and reliable in identification, and can be used for the production and identification of the Nile tilapia pseudo-female fishes.

Description

technical field [0001] The invention belongs to the field of aquatic genetics and breeding, and in particular relates to a method for highly efficient induction of pseudo-female Nile tilapia and identification of microsatellite molecular markers. Background technique [0002] There are obvious differences in the growth of tilapia females and males. In order to avoid overbreeding and take advantage of the growth advantages of male fish, the most practical and effective solution is to perform sex control on tilapia to produce male fry and breed all male fish. [0003] There are two different main sex determination systems in tilapia, and the sex of Nile tilapia and Mozambique tilapia is mainly determined by the LG1 and 23 linkage groups. [6,7] XX / XY system on LG3, while others such as blue tilapia sex are mainly determined by ZW / ZZ system on LG3 [2,3] . In addition to sex chromosomes, small factors on autosomes can also affect the main sex-determining system. However, in a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/01A01K67/027C12Q1/68C12N15/11
CPCA01K67/0275C12N15/01C12Q1/6888C12Q2600/124A01K2267/02A01K2227/40A01K2217/03
Inventor 夏军红梁明李碧君龙金华谷小慧李红莲陈朝豪
Owner SUN YAT SEN UNIV
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