The full-length cDNA sequence of glutamine synthetase from black tiger shrimps (Penaeus monodon)(denoted as PmGS) was obtained by high throughput transcriptome sequencing and RACE. On this basis, the expressions of the PmGS gene in different tissues and in hepatopancreas and gill during ammonia nitrogen stress were detected by fluorescence-quantitative real time PCR. The cDNA of PmGS was 1 420 bp,the length of the open reading frame (ORF) was 1 086 bp encoding a polypeptide of 361 amino acids,a 5'UTR of 40 bp and a 3'UTR of 294 bp. The molecular mass of the deduced amino acid (aa) sequence was 40.423kDa with an estimated pI of 6.19. And there was a poly A with 27 bp. Like other animal GSs,the structure of the PmGS protein consisted of a Glnsynt_C domain.There were eight phosphorylation sites and two glycosylation sites in this protein. Sequence alignment analysis and phylogenetic analysis showed that the PmGS with the GS of Fenneropenaeus chinensiss shared a similarity of 99.45%. Analysis of the tissue expression pattern of the PmGS showed that the PmGS mRNA was expressed in all tested tissues,including lymphoid tissue,ovary,eyestalk nerve,brain, stomach,muscle,intestines,thoracic nerve,hepatopancreas and gill. With the highest levels in lymphoid tissue, the second level in gill and the lowest level in thoracic ganglia. The expressions of the PmGS gene in hepatopancreas and gill significantly differ from the control group (P<0.05), and the expression profiles differed between hepatopancreas and gill. The result shows that PmGS might play an important role in shrimp ammonia metabolism and be involved in responses to acute ammonia stress.