Solute carrier family 11-member A1 (SLC11A1) gene encodes natural macrophage resistance-associated protein which regulates activity of macrophages against intracellular pathogens. The objective of this study was to study the polymorphism in the microsatellites present at 3′ untranslated region (UTR) of the SLC11A1 gene in 113 Zhongdian Yellow cattle (Bos taurus). Using DNA bi-directional sequencing, we detected seven alleles (GT10–16) for the first microsatellite (MS1), five alleles (GT12–16) for MS2, and four alleles (GT4–7) for MS3. MS3 is studied for the first time and revealed four novel variants (alleles GT4–7). Alleles GT12 (45.1%), GT13 (59.3%), and GT5 (85.4%) were the most frequent alleles at MS1, MS2, and MS3, respectively, Genotypes G12/12, G13/13, and G5/5 had the highest frequency 0.239, 0.540, and 0.743 at MS1, MS2, and MS3, respectively. Haplotypic data revealed that GT12/GT13 was the most frequent haplotype observed followed by GT12/14 haplotype. Three nucleotide variations were observed in MS1 and MS2. Comparative analysis of GT12/GT12 and GT13/GT13 genotype with other bovine genotypes showed significant difference (P > 0.05). Our results suggest that the homozygous genotypes GT12/GT12 and GT13/GT13 in Zhongdian Yellow cattle might be related to disease resistance. The findings reported in this study would be helpful in cattle breeding programs.