全部 标题 作者
关键词 摘要

OALib Journal期刊
ISSN: 2333-9721
费用:99美元

查看量下载量

相关文章

更多...

4种方法提取金黄色葡萄球菌基因DNA效果的比较
Comparison of Four Methods for Extracting the Genomic DNA of Staphylococcus aureus

DOI: 10.12677/HJBM.2019.94023, PP. 154-161

Keywords: 金黄色葡萄球菌,DNA提取,PCR检测
Staphylococcus aureus
, DNA Extraction, PCR Detection

Full-Text   Cite this paper   Add to My Lib

Abstract:

目的:比较4种方法,包括煮沸法、改良碱裂解法、磁珠法和吸附柱法提取金黄色葡萄球菌基因DNA效果,为临床金黄色葡萄球菌感染的快速检测提供技术支撑。方法:选取20株临床分离耐甲氧西林的金黄色葡萄球菌,分别使用煮沸法、改良碱裂解法、磁珠法和吸附柱法提取基因DNA。通过PCR法扩增所提取DNA模板中金黄色葡萄球菌spa基因、mecA基因和femB基因,以评估4种方法提取效果。结果:磁珠法制备的模板中,PCR法可以扩增出所有标本的spa基因、mecA基因和femB基因;吸附柱法制备的模板中,所有标本均可扩增出spa基因和mecA基因,而仅19个标本扩增出femB基因;以改良手工碱裂解法制备的模板,所有标本均可扩增出mecA基因,19标本可以扩增出spa基因和femB基因;煮沸法制备的模板中,PCR法可以分别扩增出7个标本的spa基因,8个标本的mecA基因,所有标本均未扩增出femB基因。结论:改良碱裂解法、吸附柱法和磁珠法提取金黄色葡萄球菌基因组DNA均可以用于PCR扩增,直接煮沸法提取效果较差,不适合用于临床金黄色葡萄球菌基因组DNA提取;磁珠法用时最短,更适合于大量快速自动化提取。
Objective: To compare the effects of four methods, including boiling method, improved alkali cracking method, magnetic bead method and adsorption column method, for extracting the genomic DNA of
S. aureus, selecting methods with better extraction quality and more suitable for automation, to provide technical support for rapid detection of clinical S. aureus infection. Methods: 20 clinical strains of methicillin-resistant S. aureus (MRSA) were isolated and their DNA was extracted by boiling method, improved alkali cracking method, magnetic

References

[1]  Pankratova, G., Hederstedt, L. and Gorton, L. (2019) Extracellular Electron Transfer Features of Gram-Positive Bacteria. Analytica Chimica Acta, 1076, 32-47.
https://doi.org/10.1016/j.aca.2019.05.007
[2]  Tateda, K., Ohno, A., Ishii, Y., et al. (2019) Investigation of the Susceptibility Trends in Japan to Fluoroquinolones and Other Antimicrobial Agents in a Nationwide Collection of Clinical Isolates: A Longitudinal Analysis from 1994 to 2016. Journal of Infection and Chemotherapy, 25, 594-604.
https://doi.org/10.1016/j.jiac.2019.03.008
[3]  Monciardini, P., Bernasconi, A., Iorio, M., et al. (2019) Antibacterial Aromatic Polyketides Incorporating the Unusual Amino Acid Enduracididine. Journal of Natural Products, 82, 35-44.
https://doi.org/10.1021/acs.jnatprod.8b00354
[4]  Gentile, A., Bakir, J., Ensinck, G., et al. (2018) Community-Acquired Methicillin-Resistant Staphylococcus aureus Infections: Hospitalization and Case Fatality Risk in 10 Pediatric Facilities in Argentina. Archivos Argentinos de Pediatría, 116, E47-E53.
https://doi.org/10.5546/aap.2018.eng.e47
[5]  Amirsoleimani, A., Brion, G.M., Diene, S.M., et al. (2019) Prevalence and Characterization of Staphylococcus aureus in Wastewater Treatment Plants by Whole Genomic Sequencing. Water Research, 158, 193-202.
https://doi.org/10.1016/j.watres.2019.04.035
[6]  Belbekhouche, S., Bousserrhine, N., Alphonse, V., et al. (2019) From Beta-Cyclodextrin Polyelectrolyte to Layer-by-Layer Self-Assembly Microcapsules: From Inhibition of Bacterial Growth to Bactericidal Effect. Food Hydrocolloid, 95, 219-227.
https://doi.org/10.1016/j.foodhyd.2019.04.037
[7]  Chauhan, J., Kwasny, S.M., Fletcher, S., et al. (2019) Optimization of a Small-Molecule Lipid II Binder. Bioorganic & Medicinal Chemistry Letters, 29, 1849-1853.
https://doi.org/10.1016/j.bmcl.2019.04.046
[8]  田小兰, 冯俊丽, 汪艺. 一种快速高效的DNA提取方法及其在qPCR检测金黄色葡萄球菌中的应用[J]. 食品科技, 2019(2): 344-350.
[9]  Du, M.H., Li, J.W., Zhao, R.X., et al. (2018) Effective Pre-Treatment Technique Based on Immune-Magnetic Separation for Rapid Detection of Trace Levels of Salmonella in Milk. Food Control, 91, 92-99.
https://doi.org/10.1016/j.foodcont.2018.03.032
[10]  Minarovicova, J., Cabicarova, T., Kaclikova, E., et al. (2018) Culture-Independent Quantification of Pathogenic Bacteria in Spices and Herbs Using Real-Time Polymerase Chain Reaction. Food Control, 83, 85-89.
https://doi.org/10.1016/j.foodcont.2016.12.025
[11]  Ernst, C., Bartel, A., Elferink, J.W., et al. (2019) Improved DNA Extraction and Purification with Magnetic Nanoparticles for the Detection of Methicillin-Resistant Staphylococcus aureus. Veterinary Microbiology, 230, 45-48.
https://doi.org/10.1016/j.vetmic.2019.01.009
[12]  Umesha, S. and Manukumar, H.M. (2018) Advanced Molecular Diagnostic Techniques for Detection of Food-Borne Pathogens: Current Applications and Future Challenges. Critical Reviews in Food Science and Nutrition, 58, 84-104.
https://doi.org/10.1080/10408398.2015.1126701
[13]  Li, J., Li, X.L., Shi, X.J., et al. (2019) A Designed Experiment: Polymorphism Analysis of Angiotensin-Converting Enzyme Gene from Human Buccal Epithelial Cells. Biochemistry and Molecular Biology Education, 47, 168-174.
https://doi.org/10.1002/bmb.21215
[14]  刘晔华, 陈锦艳, 江雁, 等. 磁珠法提取金黄色葡萄球菌基因组DNA的评价[C]//第七届中国临床微生物学大会暨微生物学与免疫学论坛. 2016: 1.
[15]  李克诚, 李琼, 夏菲, 等. 耐甲氧西林金黄色葡萄球菌spa基因分型[J]. 疾病监测, 2012, 27(11): 877-880.
[16]  黄辉, 陈颖, 安如俊, 等. MRSA中mecA及femB基因的检测与耐药相关性[J]. 微生物学杂志, 2009, 29(3): 54-56.
[17]  Nasser, A., Moradi, M., Jazireian, P., et al. (2019) Staphylococcus aureus versus Neutrophil: Scrutiny of Ancient Combat. Microbial Pathogenesis, 131, 259-269.
https://doi.org/10.1016/j.micpath.2019.04.026
[18]  Zhang, J., Zhu, M.S., Yang, Y.J., et al. (2018) Extraction of Genomic DNA via Superparamagnetic Fe3O4 Magnetic Colloidal Nanocrystal Clusters. Journal of Nanoscience and Nanotechnology, 18, 8105-8110.
https://doi.org/10.1166/jnn.2018.16399
[19]  Meng, X.Y., Yang, G.T., Li, F.L., et al. (2017) Sensitive Detection of Staphylococcus aureus with Vancomycin-Conjugated Magnetic Beads as Enrichment Carriers Combined with Flow Cytometry. ACS Applied Materials & Interfaces, 9, 21464-21472.
https://doi.org/10.1021/acsami.7b05479
[20]  Zhang, H.L., Huang, F.C., Cai, G.Z., et al. (2018) Rapid and Sensitive Detection of Escherichia coli O157:H7 Using Coaxial Channel-Based DNA Extraction and Microfluidic PCR. Journal of Dairy Science, 101, 9736-9746.
https://doi.org/10.3168/jds.2018-14730
[21]  Luo, J., Yu, J.P., Yang, H., et al. (2018) Parallel Susceptibility Testing of Bacteria through Culture-Quantitative PCR in 96-Well Plates. International Journal of Infectious Diseases, 70, 86-92.
https://doi.org/10.1016/j.ijid.2018.03.014
[22]  Wang, J., Shen, H.J., Huang, C., et al. (2017) Highly Efficient and Multidimensional Extraction of Targets from Complex Matrices Using Aptamer-Driven Recognition. Nano Research, 10, 145-156.
https://doi.org/10.1007/s12274-016-1273-9
[23]  李佳, 李亚娟, 胡俊菁, 等. 奶牛乳房炎主要致病菌DNA不同提取方法的比较[J]. 中国兽医杂志, 2017(2): 40-41.

Full-Text

Contact Us

service@oalib.com

QQ:3279437679

WhatsApp +8615387084133