Expression of Helicobacter pyloriAlpA protein and its immunogenicity

来源 :World Journal of Gastroenterology | 被引量 : 0次 | 上传用户:yingq
下载到本地 , 更方便阅读
声明 : 本文档内容版权归属内容提供方 , 如果您对本文有版权争议 , 可与客服联系进行内容授权或下架
论文部分内容阅读
AIM: To construct a recombinant strain which expresses adhesin AlpA of Helicobacter pylori (H pylori) and to study the immunogenicity of adhesin AlpA. METHODS: Gene Ab, which was amplified from H pylori chromosomal DNA by PCR technique, was sequenced and the biological information was analyzed, and inserted into the Nco I and Not I restriction fragments of the expression vector pET-22b(+) using T4 DNA ligase. The resulting plasmid pET-AlpA was transformed into competent E.coli BL21(DE3) cells using ampicillin resistance for selection. Recombinant strains were incubated in 5 mL LB with 100 μg/mL ampicillin overnight at 37 ℃. Sonication of BL21(DE3)pET-22b(+)/AlpA was analyzed by Western blot to detect AlpA immunogenicity. RESULTS: The gene encoding AlpA protein was amplified by PCR with chromosomal DNA of H pylori Sydney strain (SS1) as templates. It revealed that AlpA DNA fragment amplified by PCR had approximately 1 500 nucleotides, compatible with the previous reports. The recombinant plasmid pET-22b(+)/AB was successfully constructed. DNA sequencing showed one open reading frame with the length of 588 bp. It encoded seven conservative regions that showed good antigenicity and hydrophobicity by Parker and Welling method. Furthermore, INTERNET EXPASY, NNPREDICT and ISREC predicted that it was a porin-like structure consisting of β-pleated sheets that were embedded in the outer membrane. BLAST analyzed 836 767 protein sequences and found that the similar sequences were all belonging to H pylori OMP sequences. SDS-PAGE and scan analysis showed that the molecular weight of AB was 22.5 ku and recombinant protein amounted to 29% of the total bacterial protein, among which dissolved expression amounted to 21.9% of sonicated supernatant. The rAB purity amounted to 96% through affinity chromatography. Western blot analysis of rAB confirmed that it could be specially recognized by serum form rabbit immunized with AlpA and H pylori infected. CONCLUSION: Adhesin AlpA recombinant protein may be a potential vaccine for control and treatment of H pylori infection. AIM: To construct a recombinant strain which expresses adhesin AlpA of Helicobacter pylori (H pylori) and to study the immunogenicity of adhesin AlpA. METHODS: Gene Ab, which was amplified from H pylori chromosomal DNA by PCR technique, was sequenced and the biological information The resulting plasmid was inserted into the Nco I and Not I restriction fragments of the expression vector pET-22b (+) using T4 DNA ligase. The resulting plasmid pET-AlpA was transformed into competent E. coli BL21 (DE3) cells using ampicillin resistance Sonication of BL21 (DE3) pET-22b (+) / AlpA was analyzed by Western blot to detect AlpA immunogenicity. RESULTS: The gene was expressed in 5 mL LB with 100 μg / mL ampicillin overnight at 37 ° C. encoding AlpA protein was amplified by PCR with chromosomal DNA of H pylori Sydney strain (SS1) as templates. It revealed that AlpA DNA fragment amplified by PCR had approximately 1 500 nucleotides, compatible with the previous reports. The recombi DNA sequencing showed one open reading frame with the length of 588 bp. It encoded seven conserved regions that showed good antigenicity and hydrophobicity by Parker and Welling method. Further, INTERNET EXPASY, NNPREDICT and ISREC predicted that it was a porin-like structure consisting of β-pleated sheets that were embedded in the outer membrane. BLAST analyzed 836 767 protein sequences and found that the similar sequences were all belonging to H pylori OMP sequences. SDS-PAGE and scan analysis showed that the molecular weight of AB was 22.5 ku and recombinant protein amounted to 29% of the total bacterial protein, among which dissolved expression amounted to 21.9% of sonicated supernatant. blot analysis of rAB confirmed that it could be specially recognized by serum form rabbit immunized with AlpA and H pylori infected. CONCLUSION: Adhesin AlpA reco mbinant protein may be a potential vaccine for control and treatment of H pylori infection.
其他文献
用XPS研究了标称四价铈盐系列:硫酸铈铵、硫酸铈 、二苯基羟乙酸铈、碘酸铈、过氧化铈、二氧化铈以及三价铈盐系列:草酸亚铈、硫酸亚铈、二苯基羟乙酸亚铈、碘酸亚铈和氯化亚
为提高耐候钢使用期限,新日本制铁公司对耐候钢进行了表面处理。试验材选用耐候钢,其成分0.1%C-0.3%Cu-0.53%Cr-0.20Ni。①仅进行喷丸处理;②涂覆PVB树脂+磷酸和氧化铁等组成处理液;③除涂覆②种材料外,另涂覆丙烯
根据现行的卫生会计法规,医院购进药品的成本低于批发价成本的那部份差价,即药品的优惠价,医院做帐务处理时,一般作为“其他收入”处理,验收药品时,做如下会计分录: 借:药库
肝动脉注射THP-ADM治疗肝癌33例报告湖南省肿瘤医院内三科(410006)石珊,杨小红,罗以,王平辉,王肇炎THP-ADM(TNP-Adriamycin,吡柔比星)是一种半合成的化学结构与ADM相近的蒽环类抗肿瘤抗生素。实验研究表明其抗肿瘤活性与... T
关于取消人用狂犬病疫苗联营生产点的通知卫药发(1995)第83号卫生部长春、北京、兰州、上海、武汉、成都生物制品研究所、各生研所联合生产人用狂犬病疫苗单位:80年代末,在狂犬疫苗供应紧
“饭舵”其实就是饭勺。说起它,青年人可能会感到陌生,但对于上了一点年龄的人来说,那是再熟悉不过的餐具了。这种古老而奇特的硬木餐具长近20厘米,厚约0.5厘米,形状奇异:上
随着社会发展,人民在日常生活之中对电力的要求越来越高,生活之中的用电量也随之增加,为了满足民众对电力方面的需求,高效的电力系统必不可少,配网线路作为电力系统重要的一
2010年,漳州市关于市直国有集体企业困难职工大病医疗保险二次补偿实施以来,已享受二次补偿的困难职工共计63人,补偿金额为157462.19元。据悉,为完善漳州市医疗保障体系,帮助
在球面车床上加工球座时,由于球面在内凹台深处,用普通切削刀具加工,刀杆会与工件发生干涉,无法进行切削加工。后设计制造如图1所示的专用刀具,解决了深孔球面加工问题。刀
目的:通过观察脑梗塞大鼠神经功能及S-100β蛋白、一氧化氮(NO)的表达,探讨逐痰通络汤对急性脑梗塞大鼠的影响。方法:SD大鼠72只随机分假手术组、对照组、药物组,各组分术后1