Previous studies have identified miR169/NF-YA modules are important regulators of plant development and stress responses. Currently, reported genome sequence data offers an opportunity for global characterization of miR169 and NF-YA genes, which may provide insights into the molecular mechanisms of the miR169/NF-YA modules in maize. In our study, fourteen NF-YA transcription factors with conserved domains were identified based on maize genome loci. The miR169 gene family has 18 members that generate 10 mature products, and 8 of these mature miR169 members could target 7 of 14 ZmNF-YA genes in maize. The seven ZmNF-YA proteins were localized to the nucleus while lacked transcriptional activity. We investigated the expression patterns of the zma-miR169 members and their targeted ZmNF-YA genes in maize roots treated by drought stress (polyethylene glycol, PEG), hormone stress (abscisic acid, ABA), and salt stress (NaCl). The zma-miR169 family members were downregulated in short term (0 ∼ 48 h) and generally upregulated over the long term (15 days) in response to the three abiotic stress conditions. Most of the targeted ZmNF-YA genes exhibited a reverse correlation with zma-miR169 gene expression over both the short term and long term. Maize root elongation was promoted by PEG and ABA but repressed by NaCl over the long term. Apparently, ZmNF-YA14 expression perfectly matched the zma-miR169 expression and corresponded to root growth reversely.