Method for editing CRISPR/Cas9 genes of haliotis discus hannai

A wrinkled abalone, gene editing technology, applied to other methods of inserting foreign genetic material, using microinjection method, animal husbandry and other directions, can solve the problems of complex operation, slow development, low editing efficiency, etc. Precise editing and the effect of improving editing efficiency

Active Publication Date: 2021-12-31
INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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  • Application Information

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Problems solved by technology

[0006] However, at present, the development of gene editing technology in marine economic shellfish is still relatively slow, and it is only found in a few species, and in the only few species, the editing efficiency is still low
In abalone, only TALEN gene editing technology has been reported, but its operation is complicated and the editing efficiency is low

Method used

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  • Method for editing CRISPR/Cas9 genes of haliotis discus hannai
  • Method for editing CRISPR/Cas9 genes of haliotis discus hannai
  • Method for editing CRISPR/Cas9 genes of haliotis discus hannai

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Embodiment 1

[0042] Taking the opsin gene of the wrinkled abalone as an example, the gene editing technology of the wrinkled abalone was established, and the specific steps are as follows:

[0043] (1) Obtaining fertilized eggs of Abalone wrinkle

[0044] The parents of wrinkle plate abalone with mature gonads were selected from an abalone farm in Penglai City, Shandong Province, and artificial induction was carried out.

[0045] ①Prepare 24°C, 1000-1500D°UV seawater (the UV amount of seawater is adjusted according to the state of the female abalone. Among them, 1000D°UV seawater with a smaller UV dose can be used when the gonads of the female abalone are relatively full, and the UV dose can be used when the gonads of the female abalone are slightly poor. Larger 1500D°UV sea water);

[0046] UV irradiation dose = UV lamp power (milliwatts) × irradiation time (h) ÷ irradiated seawater volume (L)

[0047] ②Take the female abalone out of seawater at 20°C, dry it out of light for 40 minutes,...

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Abstract

The invention discloses a method for editing CRISPR / Cas9 genes of haliotis discus hannai, and belongs to the technical field of gene editing of aquatic animals. According to the method, a semi-dry fixing method is adopted during microinjection, so that not only can the haliotis discus hannai be well fixed, but also the injection efficiency is greatly improved; the optimal sgRNA and Cas9 final concentrations are adopted in microinjection, so that the survival rate and editing efficiency of the injected haliotis discus hannai larvae can be improved, and a foundation is laid for gene function research and genetic improvement of haliotis discus hannai by adopting a CRISPR / Cas9 gene editing method.

Description

technical field [0001] The invention belongs to the technical field of aquatic animal gene editing, and in particular relates to a CRISPR / Cas9 gene editing method for abalone wrinkled. Background technique [0002] Abalone is a single-shelled, herbivorous, coral reef-dwelling marine mollusc that is found in coastal areas around the world. Because of its impact on coral reef ecosystems, it is used in ecological studies and as a model animal for fertilization and developmental biology studies. Due to its high nutritional content, abalone has become an important economic shellfish for aquaculture worldwide. Wrinkled disc abalone (Haliotis discus hannai) is the main cultured species in East Asian countries. However, genome-wide selective breeding and molecular-assisted selective breeding have not been widely used in abalone genetics and breeding. The main limiting factor is that the function of abalone genes is still unclear. At present, most of the gene function studies in a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/89A01K67/033
CPCC12N15/89A01K67/0334C07K14/43504A01K2227/70A01K2267/03Y02A40/81
Inventor 张琳琳许悦吴富村
Owner INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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