Abstract The possibly higher lignin contents or altered carbon (C) allocation patterns in Bt corn hybrids, compared to their non-transgenic parental varieties, may alter the quality and quantity of plant residues incorporated into soils. In this study, we conducted a greenhouse experiment to investigate C allocation and lignin contents in Cry3Bb Bt and NonBt corn as affected by corn rootworm (CRW, Diabrotica virgifera virgifera) infestation. The partitioning of photosynthate C to various plant components was measured as short-term C allocation by a 13CO2 pulse-labeling system, and the lignin content or concentration was measured by the acid detergent method. Results showed that NonBt corn was significantly taller than Bt corn at all measured stages, likely resulting from inherent variability in the parental lines used in this study. However, there was no significant genotype effect on 13C allocation, total C and lignin content or concentration in plant tissues without CRW infestation. With CRW, the percentage of fixed 13C during labeling allocated to roots was significantly lower in NonBt than in Bt corn, likely caused by CRW damage in NonBt roots. The lignin content in NonBt roots was significantly higher with than without the CRW infestation, implying the stimulating effect of CRW possibly due to the triggered reaction of induced systemic resistance. Overall, the transgenic Cry3Bb event in MON863 corn did not affect measured variables, but CRW resistance in Bt corn affected the pattern of short-term C allocation and root lignin content compared to NonBt corn in the CRW presence, and has implications for soil C dynamics.