Essay On Engineering E. Coli Cells

The pGLO plasmid is engineered to express green fluorescent protein (GFP) in the presence of the sugar arabinose as well as the ampicillin resistance gene β-lactamase (bla) (Brown, 2011). Original E. coli HB101 do not have ampicillin resistance or the GFP gene allowing them to glow under UV light. In this experiment, we transformed E. coli HB101 with the pGLO plasmid by heat shock to make the bacterial cells competent, allowing the plasmid to enter the cell (Brown, 2011). The mixture of bacteria with pGLO plasmid were given recovery time after heat shock, then spread on LB/amp and LB/amp/ara agar plates. The bacteria mixture with no plasmid added were spread on LB and LB/amp agar plates and all four plates were incubated at 37°C for

Control plasmids lux and pUC18 were introduced into E. Coli through a process of transformation.

In this investigation pUC19 plasmids were used as the vector due to its small size of 2686bp, high uptake efficiency by the host and fast replication time. Important features of this plasmid include the origin of replication and multiple cloning sites (MCS). The origin of replication allows the plasmid to replicate inside the host bacterium. The MCS is located within the lacZ gene and contains unique sites for the Xbal & EcoRI restirction enzymes to cut and produce sticky ends for the CIH-1 gene to bind to. Furthermore, the pUC19 plasmid also contains an ampiccilin resistance gene so only transforemed E.coli are able to remain viable when spread on the agar plates that also has the addition of ampiccilin. The lacZ gene encodes the β-galactosidase enzyme which aids in indentifying the recombinant E.coli from the non recombinant cells (Coventry University 2016).

Along with this, he volunteers at a local hospital in the city, helping to improve the Calgary community. Science and biology have been an integral part of his entire life and have served as a source of inspiration. Over this past year, iGEM has provided him with the chance to further explore the field of synthetic biology along with connecting him with like minded people and peers.

The genome of E. coli 0157:H7 was recently sequenced and contains 1,387 new genes absent in non pathogenic strains. Analysis of the genome suggests that there are a variety of potential virulence genes that have yet to be explored, including fimbrial and other adhesions, secretion systems, and toxins. Many of the virulence genes are similar to those associated with pathogenesis in Salmonella and Shigella. Most of them are found on pathogenicity islands from viral DNA that is transmitted to the bacteria through the activity of a lysogenic phage. The two most important toxins, Stx1 and Stx2 (for shiga-toxin) are found on such islands and are composed of 5 B subunits surounding an active A component, which is transported into the cell after the

Based on the experiment results and the data obtained, this experiment showed that the predicted results were accurate. It is possible to produce a bacterial transformation of the genes of the bacteria E. Coli by the inoculation of a p-GLO plasmid. The bacterial transformation produced a change in the bacterial genome showing a change in the phenotype of the bacteria, by acquiring the traits of the genes introduced into the plasmid. In this experiment, each of the four plates had LB broth for nutrients and a bacterial suspension of E. Coli to study its reaction in the presence of the antibiotic ampicillin and the p-Glow plasmid. In the control plates, labeled as # I for LB/-pGLO, and # II for LB/-pGLO/

ROCKVILLE, MD and San Diego, CA (May 20, 2010)— Researchers at the J. Craig Venter Institute (JCVI), a not-for-profit genomic research organization, published results today describing the successful construction of the first self-replicating, synthetic bacterial cell. The team synthesized the 1.08 million base pair chromosome of a modified Mycoplasma mycoides genome. The synthetic cell is called Mycoplasma mycoides JCVI-syn1.0 and is the proof of principle that genomes can be designed in the computer, chemically made in the laboratory and transplanted into a recipient cell to produce a new self-replicating cell controlled only by the synthetic genome.

This experiment was performed to assess the efficacy of genetic transformations on bacteria via plasmid DNA coding for ampicillin resistance and green fluorescent protein. Genetic transformation was studied by taking transformed and untransformed Escherichia Coli (E. coli) and placing them on various media to observe gene expression via growth and color under UV light. The transformed E. coli were able to grow on ampicillin while the untransformed E. coli, which lacked the plasmid genes for ampicillin resistance, only grew on nutrient broth. In the presence of arabinose, the transformed E. coli glowed green. These results support the previous scientific understanding of bacterial competency, vectors, and gene expression and support gene transformations as an effective method to transfer the desirable DNA of one organism into another organism’s DNA. These results can be applied to real world issues such as medical treatments, food production, and environmental conservation.

Plasmid map of pRSETB. Primers were designed to amplify via PCR to cDNA. The PCR product was digested with XhoI and EcoRI enzymes and ligated into the pRSETB plasmid. The pRSETB plasmid contains a T7 promoter region, is ampicillin resistant, inducible with Isopropyl β-D-1-thiogalactopyranoside IPTG, a molecular mimic of a lactose metabolite that triggers transcription of the lac operon, and has XhoI and EcoRI cut sites. The pRSETB plasmid is transformed into dH5α E. coli and plated on carbenicillin plates. Colonies are selected and grown on a carbenicillin plate while PCR is used to check that the plasmid that was up-taken was not

The objective of this experiment was to genetically transform E.coli cells to express ampicillin resistance and to produce the green fluorescent protein by using the pGLO plasmid. It was hypothesized that only the cells with pGLO DNA added to the solution would be able to survive in the same environment as the ampicillin, and that only the cells grown in the plate with arabinose would fluoresce bright green. The results supported the hypothesis, showing that the +pGLO LB/amp/ara plate was the only plate that had fluorescent cells and grew. Additionally, that +pGLO cells were the only ones that grew on an LB/amp plate.

The experiment completed by Stanley et al, also used the transformational effectiveness of E.Coli to complete their R-Factor DNA study, so the effectiveness of using E.Coli in this type of experiment is well documented. (Stanley et al, 1972) The antibiotic that was used in our replication experiment was kanamycin, which is a amino-glycoside and it proceeds to block the areas that initiation complexes may occur. (National Center, 2017)

Radiation therapy is the ionization of atoms in tissues resulting in formation of highly reactive radicals in a well-defined, restricted volume (1). In other words, ionizing radiations are used to eradicate tumors and at the same time preserve structure and function of normal tissue. A limitation is prevented from being a problem. If bone marrow or neuronal cells are destroyed or injured, they do not regenerate. However, with radiation therapy, these cells are often saved from injury or destruction, unless the tumor is infecting bone marrow or neuronal cells. Today, radiation therapy is the most popular type of cancer therapy in use. It is used to treat one-half to two-thirds of all cancers, which translates to more than ten percent of the population

point mutation, frameshift, etc.). The advantage of using a synthetic system like ours is that the entire system is located on a plasmid and we hypothesise that the majority of mutations resulting in resistance will occur in the plasmid sequence. Unlike natural systems that contain many factors with genes encoded throughout the genome, our synthetic system allows us to monitor mutations in a more tractable manner. Further studies of the constant evolution of microecologies using this or other synthetic systems may allow researchers to tune in on “evolutionary steps” in a more accountable manner.

In 2007, it is predicted that almost 1.5 million people will be diagnosed with cancer in the United States (Pickle et al., 2007). More than half of these cancer patients will undergo the use of radiation as a means for treating cancer at some point during the course of their disease (Perez and Brady, 1998). Cancer, a disease caused by an uncontrollable growth of abnormal cells, affects millions of people around the world. Radiotherapy is one of the well known various methods used to treat cancer, where high powered rays are aimed directly at the tumor from the outside of the body as external radiation or an instrument is surgically placed inside the body producing a result of internal radiation. Radiation is delivered to the cancerous regions of the body to damage and destroy the cells in that area, terminating the rapid growth and division of the cells. Radiation therapy has been used by medicine as a treatment for cancer from the beginning of the twentieth century, with its earliest beginnings coming from the discovery of x-rays in 1895 by Wilhelm Röntgen. With the advancements in physics and computer programming, radiation had greatly evolved towards the end of the twentieth century and made the radiation treatment more effective. Radiation therapy is a curative treatment approach for cancer because it is successful in killing cancerous tumor cells and stop them from regenerating.

Bacteria play major role on production of proteins especially for proteins that are required to be synthesised in glycosylated, bacteria have outstanding appearance system and also their relative simplicity biologically. Bacterial processes also be likely to be cheaper than eukaryotic cell processes because of lesser media costs and shorter method. The most ordinarily use of bacterium for recombinant protein production is Escherichia coli, an enteric bacterium that has a long pedigree of safe use in laboratories and industry .E. coli is a mainly suitable host because it is well categorized physiologically and metabolically, it was among the first organisms to have its entire genome sequenced and many molecular biology tools a for engineering DNA sequence to generate functionally. E. coli is