All data were presented as means ± standard error of mean from at least three independent experiments. Statistical comparisons were made using Student’s t-test when comparing two groups. One-way ANOVA was performed for comparisons with more than two groups. Statistics analysis was performed using GraphPad Prism 6 software package (GraphPad Software, La Jolla, USA). In all analysis, P < 0.05 was considered to be statistically significant.
Strains are available upon request. Transcriptome data were deposited in the GEO database (accession no. GSE115806). Sup-plemental information including materials and methods, four fig-ures and four tables can be found in supplementary file. All primers used in this study are listed in Table S1.
We thank Dr. Zongbin Cui of Institute of Hydrobiology of Chinese Academy of Sciences for providing pGH-T7-zCas9 and gRNA scaffold plasmids. This work was supported by the National Natural Science Foundation of China and National Key Technologies R&D Program.
Amh, anti-Mullerian hormone; Ar, androgen receptor; Bax, Bcl2 associated X, apoptosis regulator; Bcl2, B-cell leukemia/lymphoma 2; Bmp15, bone morphogenic protein 15; Bok, Bcl2 family apoptosis regulator; Casp3a, caspase 3, apoptosis-related cysteine peptidase a; ChIP, chromatin immunoprecipitation; CRISPR, clustered regularly interspaced short palindromic repeats; Cyp19a1a, cytochrome P450, family 19, subfamily A, polypeptide 1a; DEGs, differentially expressed genes; Dmrt1, doublesex and mab-3 related transcription factor 1; ELISA, enzyme-linked immunosorbent assay; FC, fold change; FDR, false discovery rate; Fgf24, fibroblast growth factor 24; FITC, fluorescein isothiocyanate; Foxl2a, forkhead box L2a; Foxl2b, fork-head box L2b; FPKM, fragments per kilo bases per million fragments; Gapdh, glyceraldehyde-3-phosphate dehydrogenase; Gdf9, growth differentiation factor 9; GEO, Gene Expression Omnibus; GO, Gene Ontology; gRNA, guide RNA; H.E., haematoxylin and eosin; KEGG, Kyoto encyclopedia of Genes and Genomes; Mcl1, myeloid cell leukemia sequence 1, Bcl2 family apoptosis regulator; Mettl3, methyltransferase like 3; Nr0b1, nuclear receptor subfamily 0, group B, member 1; OCT, optimal cutting temperature; PI, propidium iodide; Pmaip1, phorbol-12-myristate-13-acetate-induced protein 1; QPCR, quantitative real-time PCR; Sox3, sex determining region Y-box3; Sox9a, sex-determining region Y-box 9a; Sry, sex determining region Y; Tspo, translocator protein; TUNEL, TdT-mediated dUTP nick end labeling; 17β-E2, 17β-estradiol.
Qiang Hong, Cong Li, Ruhong Ying, Heming Lin, Jingqiu Li, Yu Zhao, Hanhua Cheng and Rongjia Zhou declare that they have noconflict of interest. All institutional and national guidelines for the care and use of laboratory animals were followed.
R. Zhou and H. Cheng conceived and designed research; Q. Hong, C. Li, R. Ying, H. Lin, J. Li and Y. Zhao carried out the experimental work; Q. Hong and R. Zhou analyzed data and wrote the manuscript. All authors read and approved the final manuscript.
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