Productive tiller number (PTN) is a key agronomic trait in wheat for shaping ideal plant structure to improve grain yield. A major PTN QTL QPtn.sau-4B was localized to a 0.55 cM interval flanked by markers AX-109526283 and AX-110549715 on chromosome arm 4BS. Furthermore, we developed two pairs of near-isogenic lines (NILs, NIL-1A- and NIL-1B+; NIL-2A- and NIL-2B+) according to the allele at QPtn.sau-4B. Phenotypic analysis showed that no significant differences were detected in plant height (PH), flag leaf length (FLL), flag leaf width(FLW), spike length (SL), spike extension length (SEL) and spikelet number per spike (SNS) between NIL-1A- and NIL-1B+, except PTN and thousand kernel weight (TKW), and no significant differences were detected in PH, FLW, SL, SEL, SNS and TKW between NIL-2A- and NIL-2B+, except PTN and FLL. QPtn.sau-4B was further fine mapped in a 3.2 Mb physical interval using an F2 population. Transcriptome data analysis suggested that there were 11 up-regulated and 15 down-regulated differentially expressed genes (DEGs) associated with PTN. The KEGG pathway enrichment analysis showed that these genes were enriched in plant hormone signal transduction, carotenoid biosynthesis, cutin, suberine and wax biosynthesis and starch and sucrose metabolism. Metabolomics analysis showed that 46 credible differentially expressed metabolites (DEMs) were identified. Compared with low PTN NILs, 8 metabolites were up-regulated and 38 metabolites were down-regulated in more PTN ones. Forty-six DEMs were enriched 8 pathways, such as alpha-Linolenic acid metabolism, biosynthesis of amino acids and histidine metabolism. Functional analysis indicated that the DEGs and DEMs were related to alanine, aspartate and glutamate metabolism. These results will be useful for cloning the candidate genes and helpful for improving our understanding of the mechanisms associated with PTN development in wheat.