ABSTRACT:
INTRODUCTION:
Transformation allows for the genetic modification of a particular organism. Biotechnology is a branch of science that deals with In regard to this particular experiment, the bacteria E.coli will be placed in specific conditions under which the bacteria will be forced to take up the foreign DNA being offered for optimal survival.
METHODS:
To commence the experiment, the CaCl2 solution and the E. Coli were positioned in an ice bath. After a few minutes, without removing either container from the ice, 590 μL of the CaCl2 solution was transferred into the flask holding the E. Coli. Gently, the container with both components was then swirled to ensure the solution mixed thoroughly followed by incubation on ice for approximately 10 minutes. The incubation prepped the cells in the mixture and turned them in to competent cells, which later permitted the intake of foreign DNA (Alberte et al., 2012).
As the cells primed, two Eppendorf tubes were obtained and labeled accordingly each to represent either the control plasmid or the lux plasmid. Once labeled appropriately, both containers were placed in the ice bath tailed by the transfer of 5 μL of control plasmid and 5 μL of plasmid lux in to the pertinent tubes. Each tube then received 70 μL of the competent cells (E.coli & CaCl2). Both ampules were gently swirled and kept in ice for 15 minutes (Alberte et al., 2012).
In the mean time, an additional tube was obtained and labeled to represent no plasmids. Once labeled, 35 μL of competent cells were transported in to the container. All tube were then relocated in to a 37°C water bath where each remained for 5 minutes. After each tube had met the time requirement in the water bath, 275 μL of nutrient agar were added into the containers representing the control plasmid and the lux plasmid while 150 μL of nutrient agar was added in to the vessel representing the no plasmids. All containers were then incubated at 37°C for 45 minutes (Alberte et al., 2012).
Next, 6 agar plates were acquired and divided into two groups of three. The first three petri dishes represented the control group and thus each of the plates were labeled as follows: LB/AMP, LBC, and LB/AMPNP. The three remaining agar plates were then labeled to represent the lux plasmid as follows: LB/AMPlux, LBlux, and LBNP. 130 μL of bacterial suspension from control plasmid, 130 μL from the lux plasmid solution, and 130 μL from the no plasmid solution were then relocated in to the corresponding agar plates. Using a sterilized cell spreader for each petri dish, each solution was evenly spread across the medium. The agar plates were then covered and left to sit for about 10 minutes. The plates were then inverted and incubated at 37°C for 24 hours (Alberte et al., 2012).
RESULTS:
As indicated by Fig 1.1 & 1.2 the results obtained were not as expected. Looking at Fig 1.1, which represents the lux plasmid, it is clear that minimal growth was achieved on just one agar plate.
Fig 1.1 Lux Plasmid (a) Top agar plate labeled LB/AMPlux; No growth was exhibited (b) Bottom left agar plate labeled LB/AMPNP; No growth was exhibited (c) Bottom right agar plate labeled LBlux; Colonial growth was exhibited.
Fig 1.2 Control Plasmid (a) Top agar plate labeled LBC; No growth was exhibited (b) Bottom left agar plate labeled LB/AMP; No growth was exhibited (c) Bottom right agar plate labeled LB/AMPNP; No growth was exhibited
Discussion:
Due to the lack of observed growth it is plausible to reject predictions. It was clear through the results obtained that bacterial transformation was not attained during experimentation. This conclusion may be observed in the lack of growth is displayed in Figures 1.1 & 1.2.
Work Cited:
