Identification of sex-specific DNA markers in the army fish (Spinibarbus hollandi) by whole genome re-sequencing method

With the development of high throughput sequencing technology, genome information for more and more fish species can be available. Based on these genomes, sex-specific marker of various fish species was developed and further applied in sex control breeding. Here, whole genome re-sequencing (WGS re-seq) method was used to screen sex-specific sequences in the army fish (Spinibarbus hollandi). A total of 34.34, 30.64, 11.58, 11.21 and 12.63 Gb were generated for five females, and 12.61, 11.12, 11.67, 13.01 and 11.63 Gb were generated for five males. Moreover, we generated 39.47 Gb data for pool sequencing of female samples and 32.17 Gb for male samples. After filtering and validating, 20 candidate sex-specific sequences were obtained. Further, by PCR confirmation, six male-specific markers (X11, X17, X22, C3, C7, C10) were successfully developed with 100% accuracy in eight females and eight males. Subsequently, in another two different farmed populations of army fish, the authenticity of sex identification remained to be 100% using these markers. Besides, after homology and coverage analysis, we found the candidate sequence corresponding to C3, C7 and C10 had a similar X chromosome homologous sequence, and the resulting C3, C7 and C10 markers also amplified two bands in males but one band in females. In all, these sex-specific markers developed in this study would assist in sex control breeding and promote further research on the sex determination and differentiation mechanism of S. hollandi.