In tobacco, depending on strong shoot apical dominance, suckers grow vigorously after topping. Tobacco plants produce up to a third sucker sequentially at each leaf axil during the cultivation period. Suckers adversely affect tobacco leaf yield and quality. Therefore, sucker control is a key process to achieve tobacco production with high yields and good quality. We earlier isolated five genes involved in axial meristem formation. RNAi knock-down tobaccos of these five genes exhibited normal growth of the first sucker, but the second sucker was suppressed. The objective of this study was to evaluate the effects of these genes on sucker growth during cultivation in the field. First, knock-out (KO) tobacco mutants of each isolated gene were screened from an ethyl ethanesulfonate-mutagenized tobacco library (var. Tsukuba 1: flue-cured tobacco). Subsequently, the suckers at each leaf axil in the screened mutants were counted before and after topping in the field at the Leaf Tobacco Research Center (LTRC) in Japan. In this presentation, we present results of KO mutants of three out of five genes. The KO mutants showed normal growth of the first sucker but showed significantly suppressed second sucker formation compared with wild type. These results were similar to earlier results obtained from the RNAi knock-down tobaccos in a greenhouse. The findings imply that screened mutants may be useful for developing new varieties with fewer suckers.