Lint percentage is an important index affecting cotton fiber yield. In this study, an F2 segregation population containing 2403 individual plants was constructed derived from chromosome introgression line MBI 7747-14 (Gossypium barbadense x Gossypium hirsutum) with high lint percentage crossing with CCRI 45. The BSA-seq was conducted using two bulked samples by pooling individual plants showing higher and lower lint percentage, respectively. As a result, four candidate physical regions showing confidence indices higher than 95% were obtained in a total length of 5.47 Mb on chromosome D02, which contained 236 annotated genes. Out of them, 200 genes contain SNPs, 190 genes contain Indel, and 70 genes contain non-synonymous mutation sites. Through gene expression pattern analysis of transcriptome data, 19 candidate genes possibly associating with lint percentage were identified. GO functional enrichment analysis showed that the 19 candidate genes were enriched in NADP+ activity, aldol metabolism, carbon utilization and regulation of cell development. KEGG analysis showed that the 19 candidate genes were mainly involved in pathway such as ABC transport and phenylpropane biosynthesis. Collectively, this study laid a foundation for further analyzing the genetic mechanism of cotton fiber lint percentage formation.