The influences of electric and carbon dioxide stunning methods on ante-mortem stress and its effect on water-holding capacity of post-slaughter muscle in Duroc-Landrace-Yorkshire (with halothane genotype NN) and Sanmenxia black pigs (without halothane gene) were studied. The results showed that, compared with electric stunning, carbon dioxide stunning could significantly lower lactate (LAC), lactate dehydrogenase (LDH), creatine kinase (CK), creatinine (CRE), and cortisol (COR) content (P < 0.05) in the blood of both Duroc-Landrace-Yorkshire and Sanmenxia black pigs. The pH value measured at 45 min post-mortem (pH45min) in the longissimus muscle of pigs stunned by carbon dioxide was significantly higher (P < 0.05) than that of pigs stunned electrically. Carbon dioxide stunning significantly reduced values of muscle drip loss rate (reduced by 1.17% and 0.59%), cooking loss rate (reduced by 3.09% and 3.56%), and thawing loss rate (reduced by 0.55% and 0.29%) at 24 h post-mortem as well as shear forces (reduced by 0.71 and 0.66 kg) in the longissimus muscle of Duroc-Landrace-Yorkshire and Sanmenxia black pigs, respectively (P < 0.05). The results of low-field nuclear magnetic resonance technique (LF-NMR) indicated that carbon dioxide stunning significantly reduced the free water content (reduced by 32.24% and 28.92% for Duroc-Landrace-Yorkshire and Sanmenxia black pigs, respectively), and inhibited the transformation from immobilized to free water (state change). Thus, carbon dioxide stunning reduces the stress effect of live pigs and improves muscle water-holding capacity during pig slaughter.