Molecular breeding method for thickening intermuscular thorns of erythroculter ilishaeformis and megalobrama amblycephala
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A technology for molecular breeding of red-billed tuna, applied in biochemical equipment and methods, other methods of inserting foreign genetic materials, climate change adaptation, etc. , the effect of fast growth
Pending Publication Date: 2020-08-21
SHANGHAI OCEAN UNIV
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However, there are a large number of intermuscular spines in the muscles of red bream
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Embodiment 1
[0055] (1) Cloning of the partial nucleotide sequence of the mstn gene of the red cyper
[0056] Download the mstn nucleotide sequences of several fish species that are closely related to red bream and bream in the gene database NCBI, and perform homology comparisons, select relatively conserved regions to design PCR amplification primers, and the primer sequences are as follows: :
[0057] Cockfish mstn:
[0058] F: 5'-CATGTGGTCCAGTGGGTAATG-3' (SEQ ID NO: 3);
[0059] R: 5'-GTTGATGGGAGACATCTTGGTG-3' (SEQ ID NO: 4);
[0060] The mstn sequence was amplified by using the cDNA of the red catfish as a template, and the partial sequence of the coding region of the mstn of the red catfish was obtained, as shown in SEQ ID NO:1.
[0061] (2) Construction of the deletion mutant strain of the mstn gene of the red cyper
[0062] The gene knockout method was used to obtain the mstn gene deletion mutant line of the red cymbidium, taking the method of constructing the deletion line with...
Embodiment 2
[0087] (1) Nucleotide sequence cloning of the mstn gene of bream
[0088] Download the bream mstn sequence from the gene database, design primers, and amplify part of the CDS region of the mstn sequence. The primer sequence is:
[0089] bream mstn
[0090] F: 5'-GCAAACATCCTCTAGCACGC-3' (SEQ ID NO: 5);
[0091] R: 5'-CCACAGCGGTCTACTACCA-3' (SEQ ID NO: 6);
[0092] The mstn sequence was amplified using the cDNA of the bream as a template, and the partial sequence of the coding region of the mstn of the bream was obtained, as shown in SEQ ID NO:2.
[0093] (2) Construction of the deletion mutant strain of the mstn gene of bream
[0094] The gene knockout method was used to obtain the mstn gene deletion mutant line of the group head bream. Taking the method of constructing the deletion line with CRISPR / Cas9 as an example, the specific steps are as follows:
[0095] (i) Selection of mstn target genes in bream
[0096] The bream mstn sequence has three exons. In order to perfor...
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Abstract
The invention discloses a molecular breeding method for thickening intermuscular thorns of erythroculter ilishaeformis and megalobrama amblycephala, and belongs to the technical field of aquatic organism breeding. According to the molecular breeding method, based on a technical means of gene editing, the F0 generation of targeted mutation erythroculter ilishaeformis and megalobrama amblycephala mstn genes is obtained, and mutant individuals with mstn deletion and increased intermuscular bony area are obtained through passage. According to the invention, a parent method for obtaining erythroculter ilishaeformis and megalobrama amblycephala with increased intermuscular stabbing area by utilizing a gene editing technology is provided for the first time. The method is beneficial to large-scalecultivation of wild erythroculter ilishaeformis and megalobrama amblycephala which have thick intermuscular thorns and can be inherited in production. Being different from a transgenic method, the method can be applied to artificial breeding, overcomes the difficulty that intermuscular thorns are small and difficult to process in production, does not need to worry about the influence of transgenic food on people, is convenient for people to find the intermuscular thorns more easily when people eat erythroculter ilishaeformis and megalobrama amblycephala, and is easy to popularize in production.
Description
technical field [0001] The invention belongs to the technical field of aquatic organism breeding, and in particular relates to a molecular breeding method for thickening intermuscular spines of red bream and bream. Background technique [0002] Red bream and group-headed bream belong to the four major family fishes and are the main freshwater cultured fish in my country. Their meat is tender and delicious, and are deeply loved by consumers. They occupy a dominant position in freshwater fish farming in my country. . The use of gene editing is an important technical means for the breeding of fish traits. Through gene editing technology, specific genes can be targeted to edit specific genes to obtain new varieties with excellent traits, which is of great significance for the aquaculture industry to cultivate varieties with excellent breeding performance. At present, many researchers at home and abroad have carried out research on molecular breeding such as body color, disease r...
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