Objective To analyze the differential expression of D2 and D3 alternative splicing sites of human kinectin in human hepatocellular carcinoma (HCC) tissues, adjacent non-cancerous tissues and normal liver tissues, and to investigate a possible relationship between alternative splicing sites of kinectin and hepatocarcinogenesis. Methods The cDNA was obtained by RT-PCR in 45 coupled HCC cancerous and adjacent tissues, and 10 normal liver tissues. The difference in the expression of D2 and D3 alternative splicing sites in cDNA was examined by semi-quantitative PCR, and statistical analysis was performed. Results The ratio of D2L (long segment contains of the D2 region)/D2S (short segment that does not contain a D2 zone) in hepatocellular cancerous tissues was 2.709 ± 1.025, the ratio of D2L/D2S in adjacent non-cancerous tissues was 1.564 ± 0.357, and the ratio of D2L/D2S in normal liver tissues was 1.507 ± 0.499. The differences were statistically significant (F=29.698, P<0.05);the ratio of D2L/D2S in hepatocellular cancerous tissues was higher than that of adjacent non-cancerous tissues and normal liver tissues (P<0.05). The ratio of D3L (long segment contains of the D3 region)/D3S (short segment that does not contain a D3 zone) in hepatocellular cancerous tissues was 1.232 ± 0.041, the ratio of D3L/D3S in adjacent non-cancerous tissues was 1.156 ± 0.309, and the ratio of D3L/D3S in normal liver tissues was 1.282 ± 0.343. The ratio of D3L/D3S was not significantly different among hepatocellular cancerous tissues, adjacent non-cancerous tissues and normal liver tissues (F = 0.989, P > 0.05). Conclusion Variant containing D2 is over expressed in cancerous tissues and this alteration may be tumor associated.