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首页> 外文期刊>Research Opinions in Animal & Veterinary Sciences >Cloning and molecular characterization of omp31 gene of the Indian isolate of Brucella melitensis
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Cloning and molecular characterization of omp31 gene of the Indian isolate of Brucella melitensis

机译：印度布鲁氏菌分离株omp31基因的克隆与分子鉴定

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Brucellosis, caused by members of the genus Brucella, is an important re-emerging bacterial zoonosis and a significant cause of reproductive losses in animals. The genes encoding for the Brucella major omp31 has been extensively studied in reference and vaccine strains to determine their role in protection against infection. In this study, native major omps from field isolate of Brucella melitensis and B. abortus were extracted using N-lauryl sarcosinate. From original 2 gm (wet weight) of B. abortus and 1 gm (wet weight) of B. melitensis cells, about 5 mg (0.25%) and 2 mg (0.20%) of omp suspended in 1 ml of Tris HCl was recovered, respectively. Analysis of sonicated cells and sarcosyl extracted omps of B. abortus and B. melitensis revealed the presence of native omp31 in field isolate of B. melitensis by SDS-PAGE and Western blot, however, the omp31 gene was not present in B. abortus strain. With the exception of this, there was no any other significant difference in banding pattern and immunoreactivity between the two species both before and after sarcosyl extraction. The western blot analysis performed on native omp further confirmed that the omp31 protein is one of the major immunodominant proteins in B. melitensis. Confirmation of the absence of omp31 gene in B. abortus was also made by PCR amplification. The B. melitensis omp31 gene was PCR synthesized based on its ORF sequence and directly cloned to an entry vector. DNA sequence analysis revealed an open reading frame of 240 codons, and as compared to the consensus sequence of Brucella omp31 genes, six nucleotides have been replaced in the field isolate. The predicted sequence of omp31 showed a remarkable degree of similarity (97%) to the reported omp31 sequences of Brucella species and very less significant identity with omps from other bacteria like Yersinia enterocolitica ompH, E. coli O157 ompW, and Pseudomonas aeruginosa ompM. Being the major omp in B. melitensis and B. ovis strains, omp31 might have a particular usefulness for vaccination against sheep and goat brucellosis.
机译：由布鲁氏菌属成员引起的布鲁氏菌病，是一种重要的新出现的细菌人畜共患病，也是动物繁殖损失的重要原因。已在参考菌株和疫苗菌株中广泛研究了布鲁氏菌主要omp31的编码基因，以确定它们在预防感染中的作用。在这项研究中，使用N-月桂基肌氨酸盐提取了布鲁氏菌和流产布鲁氏菌田间分离物的天然主要omps。从最初的2 gm（湿重）流产双歧杆菌和1 gm（湿重）的B. melitensis细胞中，回收到悬浮在1 ml Tris HCl中的大约5 mg（0.25％）和2 mg（0.20％）的omp。 ， 分别。对流产双歧杆菌和肉毒双歧杆菌的超声处理的细胞和糖基提取的omps的分析显示，通过SDS-PAGE和Western印迹，在野双歧杆菌的野外分离株中存在天然omp31，但是，在流产双歧杆菌中不存在omp31基因。 。除此以外，在提取糖基之前和之后，两个物种之间的谱带模式和免疫反应性没有其他显着差异。对天然omp进行的western印迹分析进一步证实，omp31蛋白是melitensis中的主要免疫优势蛋白之一。还通过PCR扩增证实流产芽孢杆菌中不存在omp31基因。基于其ORF序列PCR合成了苜蓿芽孢杆菌omp31基因，并将其直接克隆到进入载体中。 DNA序列分析揭示了一个240个密码子的开放阅读框，并且与布鲁氏菌omp31基因的共有序列相比，在现场分离物中已替换了6个核苷酸。预测的omp31序列与报道的布鲁氏菌属物种的omp31序列显示出显着的相似度（97％），并且与来自其他细菌（如小肠结肠炎耶尔森氏菌ompH，大肠杆菌O157 ompW和铜绿假单胞菌ompM）的omps的显着性较低。作为melitensis和B. ovis菌株的主要omp，omp31可能在针对绵羊和山羊布鲁氏菌病的疫苗接种中特别有用。

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