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Application of isolated gene sequence in preparation of albino strain of Japanese oryzias latipes

A strain and sequence technology, applied in the field of medaka albino mutants in Japan, can solve the problem of low mutation frequency and achieve the effects of high mutation rate, low cost and short breeding time

Inactive Publication Date: 2018-05-11
HUAZHONG AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, medaka albino strains are all pure strains obtained through artificial screening of natural mutations. The formation of these mutants is extremely random, and the mutation frequency is very low.

Method used

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  • Application of isolated gene sequence in preparation of albino strain of Japanese oryzias latipes
  • Application of isolated gene sequence in preparation of albino strain of Japanese oryzias latipes
  • Application of isolated gene sequence in preparation of albino strain of Japanese oryzias latipes

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Embodiment 1

[0023] Application of an isolated gene sequence in the preparation of Japanese medaka albino strains:

[0024] In this example, the CRISPR / Cas9 method is used to mutate the target sequence to prepare the Japanese medaka albino strain. Other gene editing methods in the field can successfully prepare Japanese medaka albino as long as the mutation is made for SEQ ID NO.1 strain.

[0025] 1.1 Experimental materials

[0026] The wild-type Japanese medaka was bred in the fish room of the Fisheries College under the condition of 28 ℃ water temperature and 14:10 photoperiod. The embryos used in medaka microinjection were obtained from the natural spawning of male and female medaka.

[0027] 1.2 Experimental method

[0028] 1.2.1 Construction of zCas9 plasmid

[0029] The original plasmid pT3TS-nzCas9n of zCas9 was obtained from the National Zebrafish Resource Center (http: / / zfish.cn) (Jao et al., 2013). Transform pT3TS-nzCas9n into pCMV-T7zCas9 by adding the CMV-T7 promoter, incl...

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Abstract

The invention belongs to the field of molecular biology, and particularly relates to an application of an isolated gene sequence in preparation of an albino strain of Japanese oryzias latipes. The gene sequence is shown in SEQ ID No.1. Gene mutation is performed based on the gene as shown in SEQ ID No.1 as a target gene, F0 embryos with obvious pigment deletion are selected and cultured into parent fish, F1 embryos are produced by hybridization of F0 mutant parents, the F1 embryos with pigment deletion are screened out to be cultured continuously, and the stable genetic oryzias latipes albinostrain with red eyes can be produced. Compared with an oryzias latipes albino strain obtained with a traditional method, the oryzias latipes albino strain obtained with a gene mutation method has theadvantages of high mutation rate (90% or higher mutation rate), short breeding time (the mutant phenotype can be observed in 2-3 days for subsequent screening), low cost and the like, and has basic value and application value of scientific research.

Description

technical field [0001] The invention belongs to the field of molecular biology, and specifically relates to the application of an isolated gene sequence in the preparation of an albino strain of Japanese medaka. Using the sequence provided by the invention as the target sequence, a stable heredity can be obtained quickly and efficiently through phenotypic screening. albino mutant of Japanese medaka. Background technique [0002] Medaka (Oryzias latipes) has the advantages of many strains, small size, convenient breeding, strong fecundity, short reproductive cycle, obvious sex difference, known sex-determining genes, transparent embryos and fast embryonic development, etc. It is an important model organism in many fields such as environmental toxicology, cell biology and immunology (Furutani-Seiki & Wittbrodt, 2004). The genome and transcriptome data of medaka are complete (K.P.Lai et al., 2015), and embryonic stem cells, reproductive stem cells, haploid stem cells, etc. hav...

Claims

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Application Information

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IPC IPC(8): C12N15/90A01K67/027
CPCA01K67/0276A01K2217/075A01K2227/40A01K2267/02C07K14/461C12N9/22C12N15/102C12N15/902
Inventor 陈天圣方健
Owner HUAZHONG AGRI UNIV
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