Background MicroRNAs (miRNAs) play an important role in organ development and may be factors in congenital malformations.The causes of non-syndromic, isolated cleft palate (NSCP) are still unclear; the aetiology is complex and has both genetic and epigenetic factors.Method All human patients and control samples were collected from consenting individuals according to protocols approved by the ethics committee.The 50 non-syndromic, isolated cleft palate patients were from Handan Central Hospital of Hebei Province in Northern China and the smile train program in Yurnan Province.Surgeons diagnosed NSCP with the degree of NSCP was recorded from Ⅰ to Ⅲ.The Forensic Department of Fudan University collected tissue samples from 20 control subjects that were deceased as a result of traffic accidents.A total of 10 NSCP and 6 normal control samples were selected for miRNA microarray analysis.Total RNA was isolated, cDNA were synthesized and quantification of miRNAs and mRNAs were analysed by SYBRGreen qRT-PCR methods.Target genes were predicted and pathway enrichment was performed using bioinformatics analysis.Results In this study, using miRNA rnicroarray technology, we validate three over-expressed miRNAs (hsa-miR-1246, hsa-miR-1323 and hsa-miR-93) and one under-expressed miRNA (hsa-miR-143*) in patients with non-syndromic, isolated cleft palate (NSCP).In the NSCP group, hsa-miR-143* was up-regulated and the target gene was WNT5A, while hsa-rniR-1246, hsa-miR-1323 and hsa-miR-93 were down-regulated and the target gene was FGFR2.Whether miRNA expression levels are associated with an increased risk of developing NSCP was examined and correlation with age and gender were analyzed.The risk analysis identified that the odds ratios (95% CI) of hsa-miR-1246, hsa-miR-1323 and hsa-miR-93 were higher and ranged from 9.75 (1.642-57.89) to 13.67 (2.480-75.30) andp<0.05.Neither age nor gender was associated with miRNA expression levels.In males, expression levels of hsa-miR-1246, hsa-miR-1323, hsa-miR-93 and hsa-miR-143* were associated with NSCP (p value =0.0012, 0.0008, 0.0088 and 0.0385, respectively).Bioinformatics identified nine KEGG pathways enriched with predicted target genes.Conclusion In summary, rniRNAs profiles and target genes in human soft palate tissue were defined.Four distinctive miRNAs and their target genes expressed in NSCP will advance our understanding of the genetic profile of this complex disease.Further exploration will be needed to elucidate how these miRNAs and target genes rnodulate signaling pathways during palatogenesis.