宁夏地区布鲁氏菌临床分离株OMP25基因序列特征分析

目的采用PCR法扩增宁夏地区布鲁氏菌临床分离株OMP25基因并测序，分析OMP25基因序列特征，为布鲁氏菌鉴定以及基因克降与表达研究提供参考。方法参考GenBank公布的布鲁氏菌外膜蛋白OMP25基因序列设计1对引物，采用PCR法从宁夏地区分离株布鲁氏菌基因组中扩增OMP25基因并进行双向测序，测序结果用Contig Express软件拼接，采用DNAstar软件进行分析。结果经鉴定，两分离株布鲁氏菌均为羊种3型，测序获得OMP25基因全长序列为638bp，编码212个氨基酸，核苷酸序列与GenBank公布羊种布鲁氏菌OMP25基因同源性为99．9％，氨基酸序列同源性为99．7%。系统进化树显示，分离株与参考株布鲁氏菌亲缘关系相近，与羊种布鲁氏菌为同一个分枝。结论宁夏地区布鲁氏菌分离株OMP25基因序列与参考菌株序列同源性高，且与羊种布鲁氏菌存同一分枝，表明OMP25基因序列是一个保守序列。该研究结果可为布鲁氏菌的分子生物学鉴定和诊断试剂的研制提供参考。

Analysis of the OMP25 gene sequence characteristics of clinical Brucella isolates from Ningxia

Objectives To use a polymerase chain reaction （PCR） to amplify the OMP25 gene from clinical Brucella iso lates from Ningxia and analyze the characteristics of the OMP25 gene sequence in order to provide a basis for Brucella i dentification and gene cloning and expression studies. Methods A pair of primers for the gene sequence of OMP25 of Brucella was designed after referring to C-enBank. PCR was used to amplify the omp25 gene from the Brucella genome, and gene fragments were detected using agarose gel electrophoresis. PCR products were sequenced and assembled into a single sequence using Contig Express software. Sequencing results were analyzed using DNAstar software. Results Brucella strains were identified as Brucella melitensis type 3, and sequencing resulted in a full-length sequence of the （）MP25 gene of 638 bp. This sequence encodes 253 amino acids and has similarity of 99.9% to the nucleotide sequence of the OMP25 gene of B. re~elitensis published in GenBank. The similarity of amino acid sequences was 99.7%. A phylogenetic tree revealed that isolates were closely related to the reference strain of Brucella, and B. melitensis was one branch of that tree. Conclusion The OMP25 gene of Brucella isolates was homologous to that of Brucella reference strains and B. melitensis was one branch of Brucella＇s phylogenetic tree. The OMP25 gene is a conserved sequence. The current results provide a basis for molecular biological identification of Brucella and preparation of diagnostic reagents to detect Brucella.