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In addition, disruption of Dmrt1 in XY fish resulted in increased expression and serum estradiol-17β and 11-ketotestosterone levels.On the contrary, deficiency of Foxl2 in XX fish exhibited varying degrees of oocyte degeneration and significantly decreased aromatase gene expression and serum estradiol-17β levels.
Gene targeting has never been easy for other organisms.
Six pairs of TALENs were constructed to target genes related to sex differentiation, including mutation on gonadal phenotype, sex differentiation, and related gene expression were analyzed by histology, immunohistochemistry, and real-time PCR.
In Dmrt1-deficient testes, phenotypes of significant testicular regression, including deformed efferent ducts, degenerated spermatogonia or even a complete loss of germ cells, and proliferation of steroidogenic cells, were observed.
The DNA binding domain consists of a variable number of tandem repeats of a 34-amino-acid monomer, which specifies the DNA binding sequence by its 12th and 13th repeat-variable di-residues (RVDs, which specifically recognize a single nucleotide) (8, 9).
It is known that the RVDs NI, NG, HD, and NN preferentially recognize adenine (A), thymine (T), cytosine (C), and guanine (G)/adenine (A), respectively (10).
These double-strand DNA breaks are frequently repaired through nonhomologous end joining, introducing indels (insertion or deletion) that frequently result in frameshift mutations (8, 9).
The DNA binding specificity of TALENs, as distinct from ZFNs, is based on the transcription activator-like effectors (TALEs) from plant pathogens, which contain variable N and C termini but a conserved central domain for specific DNA binding.Taken together, our data demonstrated that TALENs are an effective tool for targeted gene editing in tilapia genome.Foxl2 and Dmrt1 play antagonistic roles in sex differentiation in Nile tilapia via regulating Reverse genetics approaches have been important in demonstrating gene functions, genetic engineering, and understanding complex biological processes.Up to now, no direct evidence has been provided to clarify the effects of Dmrt1 and Foxl2 knockout on sex differentiation of tilapia and other teleosts.In this report, gene targeting was successfully conducted in tilapia using TALENs.Therefore, it is urgent to establish an efficient and precise genome editing method to study the functional genomics of Nile tilapia.