Vaccination with live attenuated vaccines (LAVs) is an effective way for prevention of infectious disease. While several methods are employed to create them， efficacy and safety are still a challenge. In this study， we evaluated the feasibility of creating a self-attenuated RNA virus expressing a functional species-specific artificial microRNA. Using influenza virus as a model， we produced an attenuated virus carrying a mammalian-specific miR-93 expression cassette that expresses a viral nucleoprotein (NP)-specific artificial microRNA from an insertion site within the non-structural (NS) gene segment. The resulting engineered live-attenuated influenza virus， PR8-amiR-93NP， produced mature and functional artificial microRNA against NP in mammalian cells， but not in avian cells. Furthermore， PR8-amiR-93NP was attenuated by 10 fold in mice compared with its wild-type counterpart. Importantly， intranasal immunization with PR8-amiR-93NP conferred cross-protective immunity against heterologous influenza virus strains. In short， this method provides a safe and effective platform for creation of live attenuated RNA viral vaccines.