Aim To construct and identificate recombinant adenovirus with siNrdp1 gene using AdMax system, and investigate the effect of Nrdp1 gene on cardiomyocyte hypertrophy. Methods Designing and synthesizing siRNA sequences targeting of Nrdp1 DNA, then cloned into the shuttle vector GV119 and homologous recombinated with adenovirus backbone plasmid AdMax in BJ5183 bacteria transfected HEK293 cells, and got adenovirus containing Nrdp1-siRNA gene through packaging. Real-time quantitative PCR and Western blot were used to detect Nrdp1 expression in primary rat neonatal cardiomyocytes. After adenoviral containing siNrdp1 transfection and angiotensin Ⅱ （AngⅡ） stimulation, real-time quantitative PCR was used to detect the expression of myocardial hypertrophy marker gene （ANF, β-MHC and Skeletal-α-actin） of rat neonatal cardiomyocytes. Results Digested PCR analysis and sequencing showed that interference Nrdp1 adenovirus was successfully constructed, and the titer of virus was 1.5E＋9 PFU/mL. Real-time PCR and Western blot indicated that the expressions of Nrdp1 mRNA and protein were greatly inhibited after infection in rat primary cardiomyocytes with recombinant adenovirus particles （P<0.001）. Nrdp1 gene silencing cloud significantly increase expression of AngⅡ induced cardiomyocyte hypertrophy marker genes including ANF, β-MHC and Skeletal-α-actin （P<0.01）. Conclusion The recombinant adenovirus vector containing the Nrdp1-siRNA gene was successfully constructed, which can effectively silence Nrdp1 gene and enhance AngⅡ induced cardiomyocytes hypertrophy in vitro.