Lb-Pglo Lab Report

The plate with the bacteria that is most like the original non-transformed E. coli colonies is the LB/-pGLO plate. The bacteria could grow because there was no ampicillin on the plate, and it couldn’t have been transformed because no pGLO was added. Since the plasmid wasn’t added and the environment was ampicillin free, the bacteria on the LB/-pGLO plate should be pretty much the same as the original E. coli because nothing was done to it.

In preparing for the bacterial transformation, DNA plasmid is introduced into the E. coli cells that will express newly acquired genes. Two tubes were used and labeled both as +pGLO and -pGLO. A solution of (CaCl2) was transferred 250 µl onto the two tubes. The tubes were placed on the ice. A sterile loop was then used to gather a single colony of bacteria from a starter plate. Now, that both tubes contain bacteria they were placed on the ice for 10 minutes. Four agar plates were labeled as: +pGLO LB/amp, +pGLO LB/amp/ara, +pGLO LB, -PGLO LB/amp. Heat shock was used to transfer both the +pGLO and -pGLO, at exactly 42°C. Time was observed for 50 seconds and quickly return the tubes to the ice for another 2 minutes. As the tubes, cold down they

The focus of this lab was to identify which plasmid (PFG or Pglo) was inserted into the E. coli culture. In order for the E. coli to be transformed, the E.coli must first be made competent. The cells were made competent by incubating the cells in calcium (2+) and then in cold temperatures. A quick heat shot opens pores in the cell membrane and allowed plasmid DNA to enter the cells. The plasmid that was inserted into the E. coli culture had antibiotic-reistance gene and a reporter gene. A reporter gene is a gene that is easy to detect, and for this lab it could be Pglo or PFG. The cells were placed in plates that contained antibiotics to kill any bacteria that did not uptake the plasmid, this is necciary because about 1% of bacteria will be transformed. This allowed only cells that up took the DNA to

While leading this analysis, gene expression was critical to the results of the non-transformed and changed cells. In its procedure of choice, the bacterial cells should have picked if they ought to allow the ampicillin safe quality, therefore indicating commendable components of quality expression. The Escherichia coli's cells were not changed, there seems to have been an error during the experiment. There should have been a slight change in a way that there should have been a colonial growth design in the bacterial cells that had ampicillin safe plasmid. (Hidalgo, Nuria Fernández) On the other hand the cells that were on the ampicillin (lux, control, and NP) demonstrated a colonial design because of the non-transformed cells dying due to

Introduction Genetic transformation is known to be the conversion of a genotype into another with the addition of DNA from an external source. Frederick Griffith was the first person to use genetic transformation in 1928 with Streptococcus pneumoniae (Griffiths et al. 2000). There are three different methods that are used in genetic transformation to get the vector DNA into the nucleus of the competent cell. The methods are, projectile bombardment, electroporation, and heat shock. Heat shock is the most common method is projectile bombardment because it guarantees the mixture of the two DNA (APSnet…2014).

Bacterial transformation is one of the main ways scientists are genetically engineering cells. The reason why bacteria is getting used for these experiments is because bacteria’s genes are very versatile. For the purpose of this experiment we used the bacteria Escherichia Coli, the reason for this is that this specific bacterium is resistant to antibiotics. Plasmids were used in this experiment for the purpose of identifying what type of growth will occur also because plasmids are needed in order for transformation to be possible. Plasmid is not the only thing needed, also needed are vectors and a host. The bacteria used, E-Coli, served as the host in this experiment and the plasmids served as the vectors. Vectors are used to deliver foreign

Transformation in bacteria is something that could be essential for survival in a bacteria. In order to perform this transformation naturally a bacterium must considered competent, otherwise it must undergo an artificial transformation. Being a competent cell means that the bacteria can take up DNA from its environment naturally (5). Those that are not competent such as Escherichia coli that are not naturally competent can be tested with an artificial transformation, such as what we will use in this experiment. Methods used can obtain things such as chemical mutagens or radiation (1). The gene used for the transfer is the GFP gene (Green Fluorescent Protein), which gives an illuminating appearance under a UV light when conducted properly,

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Transformation was discovered by Frederick Griffith in 1928 in his classic experiment on Streptococcus pneumoniae. Unknown to him, his “transforming factor” which changed the non-pathogenic strains into pathogens was DNA. This was discovered in 1944 by Avery, MacLeod, and McCarty who, using isolated DNA from S. pneumoniae, were able to again cause virulence in a previously harmless strain.

In 1928, Frederick Griffith discovered transformation when observing that living cultures of a non-pathogenic strain called a Streptococcus pneumonia killed mice after being mixed with a heat-killed pathogenic strain. Griffith named concluded this as transformation after the non-pathogenic strain transformed into a pathogenic strain. In 1944, Oswald Avery purified RNA, DNA, and protein from a virulent strain of bacteria. His experiments not only revealed how virulence the transformation from S. pneumonia was but, led to the recognition of DNA as a genetic material. The exact mode of transformation varies from one bacterial species to the next. DNA is transferred between bacteria using two main methods: transmission and conjunction. In transformation,

coli cells were transformed using chemical method and were then spread-plated onto LB plates supplemented with Kanamycin 30 μg/mL (Sigma), Isopropyl β-D-1thiogalactopyranoside (IPTG) 0.5 mM (Progene) and 5-bromo-40chloro-3-indolyl-β-D-galactopyranoside (X-Gal) 80 μg/mL (Promega) and incubated overnight at 37°C.

The purpose of this literary review is to discuss the current research of biotechnology specifically bacterial transformation. Bacterial transformation is when cells take up and recombine strands of DNA (Redfield, 1993). Bacterial transformation has been thought to be the simplest process of evolving genetic exchange (Redfield, 1993). Bacterial transformation has opened research opportunities for many people.

DNA encodes the genetic instructions for cells to carry out their daily activities. DNA can come in many forms; plasmids for example are small circular DNA molecules found in most bacterial cells. Though plasmids may not be essential for the life of bacteria, it can give cells resistance in foreign environments. For the purpose of this experiment, an ampicillin-resistant plasmid is introduced to E. coli. This is done through a process of genetic engineering called transformation. Transformation works through the uptake, incorporation, and expression of a foreign gene to alter the genetic code of a cell. Three conditions are needed for successful transformation: a host, a vector, and a technique to identify the transformed cells. E. coli is used in this experiment as the host (E. coli is commonly used in biotechnology due to its rapid rate of growth and short reproduction time). A vector mediates the transfer of foreign DNA into the host cell. Plasmids are commonly used vectors that will also be used in this experiment. The procedure of tagging is used in this experiment to differentiate the transformed cells from those that were not. The learning objectives of this experiment are to: observe the process of bacterial transformation in an experiment; and demonstrate a change in phenotype due to uptake and expression of the genes in a known plasmid.

Bacterial transformation plays a huge role in this lab. Bacterial transformation is the process by which bacterial cells take up naked DNA molecules. If the foreign DNA has an origin of replication recognized by the host cell DNA polymerase, the bacteria will replicate the foreign DNA along with their own DNA. The field of study that uses bacterial transformation as their driving force for research is biotechnology. The lab that was carried out in class is just a very small example of biotechnology. Biotechnology according to www.bio.org is just simply technology based on biology that harnesses cellular and bimolecular processes to develop technologies and products that help improve our lives and the health of our planet. This field of study is used all over the world to provide for solutions for many problems that previously were not able to be solved. Some examples of areas where this technology might be used, range from the medical field, all the way to the other side of the spectrum in the agriculture business. Many patients are given the opportunity to be treated with different kinds of medicines that would not be possible without biotechnology. An example of this would be human insulin. Some people are not able to make their own insulin, this disease is called diabetes. The human insulin gene is injected into these bacteria and as a result these bacteria make human insulin which is then harvested and sold for the people that can’t make

The transformation of competent bacteria cells is to be used to take up DNA sequence of interest.