唾液链球菌尿素酶基因ureIABCEFGD活性表达与镍离子的关系

上海口腔医学 ›› 2013, Vol. 22 ›› Issue (5): 504-507.

唾液链球菌尿素酶基因ureIABCEFGD活性表达与镍离子的关系

王艳¹，李存荣²，陶丹英¹，冯希平¹

(1.上海交通大学医学院附属第九人民医院?口腔医学院口腔预防科，上海市口腔医学重点实验室，上海 200011；2.上海市口腔病防治院口腔预防保健处，上海 200001)

收稿日期:2013-03-18 修回日期:2013-04-26 出版日期:2013-10-10 发布日期:2013-10-10
通讯作者: 冯希平，Tel: 021-33183424，E-mail: fxiping1808@qq.com
作者简介:王艳(1978-)，女，博士，主治医师，现工作单位为上海市口腔病防治院口腔预防保健处，E-mail: serene2000@163.com
基金资助:
上海市自然科学基金(08ZR1416800)；上海市科学技术委员会资助项目(11411950900)

Relationship between ureolytic activity expression of Streptococcus salivarius urease genes ureIABCEFGD in Escherichia coli and nickel ions

WANG Yan¹, LI Cun-rong ², TAO Dan-ying ¹, FENG Xi-ping¹

1.Department of Preventive Dentistry, Ninth People’s Hospital, College of Stomatology, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology. Shanghai 200011; 2.Department of Preventive Dentistry, Shanghai Municipal Hospital for Oral Health. Shanghai 200001, China

Received:2013-03-18 Revised:2013-04-26 Online:2013-10-10 Published:2013-10-10
Supported by:
Supported by Natural Science Foundation of Shanghai Municipality(08ZR1416800) and Research Fund of Science and Technology Committee of Shanghai Municipality(11411950900).

摘要/Abstract

摘要： 目的：克隆唾液链球菌57.Ⅰ的尿素酶基因ureIABCEFGD转化大肠杆菌，检测重组菌株的尿素分解活性与外源性镍离子浓度的关系。方法：将目的基因分成前、后2段分别克隆，再酶切、连接并测序鉴定，得到的尿素酶基因ureIABCEFGD质粒转化感受态大肠杆菌TG-1，分别添加不同浓度的NiCl2，经Nessler试剂盒检测其分解尿素的产氨量，采用SPSS17.0软件包对产氨量和NiCl2浓度进行线性相关分析。结果：克隆的尿素酶基因ureIABCEFGD序列正确，该克隆转化大肠杆菌后，随着外源性NiCl2浓度增高，重组菌株的产氨量迅速增高，呈正相关(r=0.9714，P<0.01)；当NiCl2浓度增加到50 μmol/L时，产氨量趋于峰值，不再随NiCl2的浓度而增加。结论：唾液链球菌尿素酶基因ureIABCEFGD尿素分解活性的表达在一定浓度范围内与外源性镍离子的添加呈正相关，该克隆可用于进一步研究尿素分解活性的调控机制和方法。

关键词: 链球菌, 尿素酶, 镍, 龋病

Abstract: PURPOSE: To obtain the clone of Streptococcus salivarius 57.Ⅰ urease genes ureIABCEFGD and investigate the relationship between ureolytic activity expression of this clone in Escherichia coli and nickel ions. METHODS: The target gene was cloned by polymerase chain reaction in 2 parts separately. Then, 2 plasmids were digested by specific restriction enzymes and ligated together. The obtained plasmids were subjected to nucleotide sequence analysis and transformed into E.coli TG-1. The recombinant E.coli was added without or with different level of NiCl2. The amount of ammonia generated by ureolytic activity of each sample was measured by Nessler’s assay. SPSS 17.0 software package was used for correlation analysis. RESULTS: The clone of urease genes ureIABCEFGD was proved by sequence analysis and BLAST search. The amount of ammonia generated by the recombinant strain had a positive correlation with the level of NiCl2(r=0.9714，P<0.01). When the level of NiCl2 was 50 μmol/L, the amount of ammonia reached maximum and would have little variance despite the increase of NiCl2 level. CONCLUSIONS: Ureolytic activity expression of ureIABCEFGD has a positive correlation with the level of added NiCl2 not exceeding 50 μmol/L. The present clone can be used to further investigate the regulation of the ureolytic activity expression of Streptococcus salivarius urease gene.

Key words: Streptococcus, Urease, Nickel, Dental caries

中图分类号:

R780.2

王艳, 李存荣, 陶丹英, 冯希平. 唾液链球菌尿素酶基因ureIABCEFGD活性表达与镍离子的关系[J]. 上海口腔医学, 2013, 22(5): 504-507.

WANG Yan, LI Cun-rong , TAO Dan-ying , FENG Xi-ping. Relationship between ureolytic activity expression of Streptococcus salivarius urease genes ureIABCEFGD in Escherichia coli and nickel ions[J]. Shanghai Journal of Stomatology, 2013, 22(5): 504-507.

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