Bacteria Transformation in Biotechnology
Abstract Some bacteria are able to go through transformation making new combinations of genes. Transformation is a way of gene variability in bacteria. This experiment is based on the transformation mechanism of bacteria and gene regulation. The bacteria used for the experiment was Escherichia coli and the genes introduces for the transformation were: gfp and bla by a pGLO™ plasmid. After the insertion of the target genes and growing the bacteria on specialized LB media, it could be seen that the transformants were positive for the gene expression. The transformed E. coli on the media appeared fluorescent green under UV light.
Introduction The bacteria used in this experiment is Escherichia coli which is not naturally competent. E. coli is a gram negative rod shaped bacteria and a facultative anaerobe. This bacteria forms part of the bacterial flora in the human intestine tract. The competence of a bacteria is based on its ability to take up naked DNA from the environment and incorporated on theirs, transformation. Alteration in the permeability of the membranes allows DNA to cross the cell envelope of E. coli. Since the outer membrane of the E. coli is mostly negatively charged and the DNA molecule also has a negative charge, then the addition of CaCl2 will neutralize the interaction so that the naked DNA molecule can enter the cell. (Microbe Library web) Another important factor on the competence of the bacteria is a procedure of alternating temperature between ice bucket and heat shocks. By the combination of this two procedures E. coli becomes competent. This procedure was first reported by Mandel and Higa. (Singh 562) Even though it works it is only believed that CaCl2 helps DNA absorption to cell surface and the heat-shock step facilitates penetration of absorbed DNA into cell. (Panja 411) The main purpose of this experiment is to transform the bacteria to make it resistant to the antibiotic ampicillin. A secondary transformation is being made, and
Introduction The bacteria used in this experiment is Escherichia coli which is not naturally competent. E. coli is a gram negative rod shaped bacteria and a facultative anaerobe. This bacteria forms part of the bacterial flora in the human intestine tract. The competence of a bacteria is based on its ability to take up naked DNA from the environment and incorporated on theirs, transformation. Alteration in the permeability of the membranes allows DNA to cross the cell envelope of E. coli. Since the outer membrane of the E. coli is mostly negatively charged and the DNA molecule also has a negative charge, then the addition of CaCl2 will neutralize the interaction so that the naked DNA molecule can enter the cell. (Microbe Library web) Another important factor on the competence of the bacteria is a procedure of alternating temperature between ice bucket and heat shocks. By the combination of this two procedures E. coli becomes competent. This procedure was first reported by Mandel and Higa. (Singh 562) Even though it works it is only believed that CaCl2 helps DNA absorption to cell surface and the heat-shock step facilitates penetration of absorbed DNA into cell. (Panja 411) The main purpose of this experiment is to transform the bacteria to make it resistant to the antibiotic ampicillin. A secondary transformation is being made, and
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