Xylans are the principle non-starch polysaccharides of wheat in poultry diets which can increase the intestinal viscosity and decrease the nutrient absorption. Xylanases catalyze the hydrolysis of xylans. The aim of present study was to clone xylanase gene in pGEM vector and sequencing of this gene from the Bacillus subtilis. Genomic DNA from B. subtilis was isolated and amplified by PCR using Xylanase specific PCR primers. Then xylanase was cloned by T/A cloning technique and transformed into TOP10 E. coli cells. Finally, after sequencing, xylanase sequence similarity was checked using nucleotide BLAST analysis. The results of present study showed that xylanase was successfully cloned in pGEM vector. Sequencing confirmed that xylanase was cloned and the length of xylanase was 661 bp. BLAST search showed that the sequence of xylanase gene of the B. subtilis has 99% homology with other records existing in GenBank.