The invention provides a RanBP9 mutant gene suffering from c.126C>T mutation. The nucleotide sequence of the RanBP9 mutant gene is shown in a SEQ ID NO:1, and the RanBP9 mutant gene is a virulence gene of human male dyszoospermia. The invention further provides a RanBP9 virulence gene exon mutant screening method based on combination of multi-PCR capture and next-generation DNA sequencing. According to the RanBP9 virulence gene exon mutant screening method, finally, Sanger sequencing is adopted to verify mutation. PCR capture primer groups include amplified primers of which the sequences are shown in SEQ ID NO:7-12, and the sequences of amplified primers, used for Sanger sequencing, of a RanBP9 gene c.126C>T mutant fragment are shown in SEQ ID NO:7-8. Forms of the RanBP9 virulence gene arereported for the first time, and bases can be provided for parsing the pathogenesis of the human male dyszoospermia, establishing a detection method of the human male dyszoospermia and the like.