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:: Volume 6, Issue 1 (1-2019) ::
JBRMS 2019, 6(1): 49-56 Back to browse issues page
Investigating the effect of thermal shock factors and different cell treatments on bacterial transformation efficiency
Ciamak Ghazaei
Department of Microbiology, University of Mohaghegh Ardabili, Ardabil, Iran , ciamakghazaei@yahoo.com
Abstract:   (152 Views)
Introduction: Molecular transformation of bacterial cells plays a central role in molecular DNA transfer. At present, transformation is carried out either by chemical methods or by electric shock (electroporation). This study tended to investigate the effect of thermal shock factors and different cell treatments on transformation efficiency in E.coli.  
Materials and methods: In this method, E.coli cells were treated by different concentrations of peptide CM11 (0.1, 1, 2, 3 and μg/ml) and lysozyme enzyme (40 mg/ml and 80 mg/ml) based on competence by calcium chloride (100 and 200 mM). They also underwent thermal and cold shocks at different temperature intervals. Then, pUC19 plasmid (2.7 kbp) was transferred as a model and separately to the competent bacteria.
Results: The results showed that increasing the thermal shock in three temperature intervals (37, 42 and 47°C) with cold shock (0 and 5°C) led to an increase in the number of transformed colonies and it has a significant relationship with transformation (P<0.05). Increased CaCl2 concentration was a positive factor in the amount of transformation, but no significant relationship was found between increased lysozyme concentration and transformation (P>0.05). Increased CM11 peptide concentration showed a significant relationship with transformation (P<0.05). There was no significant relationship between three transformations (P>0.05), while there was a significant relationship between peptide CM11 concentration and lysozyme enzyme concentration in transformation simultaneously (P˂0.05).
Conclusion: It can be concluded that increased thermal shock and stability of cold shock at minimum temperature (0°C), as well as increased CaCl2 concentration and increased peptide concentration to 1 μg/ml can increase the number of transformed colonies in E.coli. Finally, peptide concentration and enzyme concentration are introduced as a simple and convenient method for increasing the transformation efficiency.
Keywords: Transformation, Escherichia coli, Plasmid, Lysozyme Enzyme, Peptide CM11
Full-Text [PDF 871 kb]   (38 Downloads)    
Type of Study: Research | Subject: Microbiology
Received: 2018/08/19 | Accepted: 2018/10/21
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Ghazaei C. Investigating the effect of thermal shock factors and different cell treatments on bacterial transformation efficiency. JBRMS. 2019; 6 (1) :49-56
URL: http://jbrms.medilam.ac.ir/article-1-389-en.html


Volume 6, Issue 1 (1-2019) Back to browse issues page
مجله ی تحقیقات پایه در علوم پزشکی Journal of Basic Research in Medical Sciences
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