To establish the mouse-lethal model for pandemic H1N1 influenza virus, provide an animal model for studying the pathogenicity and host adaptation of 2009 pandemic H1N1 influenza virus, and find out the key amino acid mutations which may affect viral virulence and replication. A pandemic H1N1 influenza virus strain, A/Sichuan/SWL1/2009 (H1N1, SC/1) was passaged in mouse lung by 15 cycles with intranasal infection. The passaged viruses were all propagated in MDCK cells and sequenced. Based on the sequencing results, four mice in each group were inoculated with 6 selected viruses and their weight and survival rate were monitored during the following 14 days after infection. Additionally, SC/1-MA P14 and P15 viruses were sequenced after purification by Plague Assay. Viral virulence was increased after serial passages and the mortality of 100% was detected after 7 passages. Several amino acid residue mutations of passaged viruses which may contribute to the enhanced virulence were observed. The increased virulence of passaged viruses and mammalian host adaptation maybe associated with amino acid mutations in viral functional proteins. Finally, we established a mouse-lethal model.