ObjectiveTo establish an animal model of TNNT3((R69H)) mutation transgenic mice. MethodsTransgenic component pEGFP-TNNT3(R69H) was constructed, linearized and purified, then microinjected into fertilized mouse eggs. These eggs were transplanted into pseudopregant mice. The genotype of transgenic founders were identified by PCR and Southern blot. The expression of TNNT3(R69H) transcript in the tissues of the transgenic mice was detected by RT-PCR and Western blot. ResultsTransgenic BDF1 mice were obtained by microinjection. PCR and Southern blot results showed 5 of 40 mice were integrated. ConclusionsTNNT3((R69H)) mutation transgenic mouse models are successfully established by pronuclear microinjection.