Cytidine functions as an essential precursor for nutraceuticals and pharmaceuticals. The global dual-function transcription factor Cra (catabolite repressor/activator) modulates carbon flux distribution during cytidine synthesis in Escherichia coli. Here, we examine how Cra regulates carbon partitioning and gene expression to influence cytidine production. We generated a cra knockout strain (E. coli NXBG-18C) from E. coli K-12 using CRISPR-Cas9 and compared it with the wild-type strain through transcriptomics (RNA-Seq) and metabolic flux analysis (¹³C-MFA). Cra deletion substantially increased cytidine yield without impairing growth, achieving a 3.77-fold higher titer (9.55 ± 0.29 g/L), a 1.21-fold elevated glucose consumption rate, and a 3.13-fold improvement in yield. Transcriptomic analysis revealed 1,819 differentially expressed genes (1,043 upregulated, 845 downregulated), with upregulated genes predominantly involved in ribosome biogenesis and the TCA cycle, while downregulated genes were linked to purine, fructose, mannose metabolism, and pentose/glucuronate interconversions. Notably, glucose PTS genes were upregulated, whereas cytidine salvage pathway genes were suppressed. ¹³C-MFA demonstrated metabolic flux redistribution, with carbon flow favoring the TCA cycle during the log phase (8h), elevating NADPH, ATP, and PRPP levels, while enhancing flux through central carbon pathways (EMP, PPP, TCA) and energy-related metabolites during the pre-production phase (16h). These findings indicate that Cra deletion reprograms central carbon metabolism by upregulating glucose uptake and TCA cycle activity, downregulating competing pathways and cytidine salvage, and ultimately redirecting carbon and energy resources toward cytidine synthesis. Cra emerges as a pivotal regulator of cytidine production through its influence on central carbon metabolic genes.