the gene expression

The bacterial transformation has been widely used in the development of different medicines and useful compounds. It lays the groundwork for the advancement of bacterial medicine by allowing the growth and action of selected bacterial species to be studied under controlled conditions. The whole idea of bacterial transition stems from the fact that genetic codes (DNA & RNA) can be transferred from one organism to another. The end product seems to be a superior creature with a plethora of attractive qualities and features (Bruijn 53). In light of the above, the study employed E. colias a test subject. The whole process involved exposing the bacteria to a gene that has the capabilities of withstanding an antibiotic known as ampicillin. Calcium chloride was a vital reagent as per the study since it allowed for the cell pores to be opened and the actual entrance of the DNA into the organism. Heat shock was also employed. For this purpose, calcium chloride was used to break the repulsion that is existent between the cell wall and DNA. The findings revealed that there was a growth of colonies in theLBC and LBNP culture plates while no development was noted in the LB/AmpNP, LB/AmpC, LB/Amplux plates. This shows that ampicillin has the ability to restrict division hence the growth of colonies.

Introduction

DNA is the storehouse for genetic material in living organisms. Evolution is guaranteed by the existence of superior genes that serve to ensure the organism can withstand conditions of the environment. Today’s advancements in technology and research have shown that it is possible to transfer genetic information from DNA to RNA through a series of intricate and well-defined processes. This act is referred to as transcription. According to Brujin, (201) Transcription is the process by which the information needed for protein synthesis is copied to mRNA once the transfer is complete, the mRNA can leave the nucleus and attach itself to a ribosome which initiates the protein synthesis process. It is worthwhile to note that transcription takes place very differently on prokaryotes since they have an apparent absence of a nucleus(Huang et al., 104). The chromosomes that are storehouses for genetic material (DNA & RNA) are found in the nucleus. Chromatin that contains genetic material is also located in there. The absence of nuclei in prokaryotic organisms causes DNA to exist in circular structures referred to as Plasmids. Plasmids are small DNA molecules with an elliptic shape (circular shape).

The primary role of these plasmids is to increase bacteria survival rate in an environment that would have been unconducive for growth and spread. This is mainly because these plasmids contain genetic material that is subject to mutation hence resistance. In light of the above, two plasmids (pUC18 and lux) will be used(Bullerwell, 67). It is important to note that the plasmids being used will be in possession of the ampicillin resistant gene. Control of gene expression involves the overall binding of transcription factors to DNA. The transcription factors are therefore referred to as specialized proteins. The role of these transcription factors includes preventing and fostering the transcription of specific genes. The overall process is so organized such that the elements can ensure only the right number of genes is able to enter into the organism in question. The transcription factors are able to control the process because they are protein in nature. In light of the above, transcription is determined by a variety of factors and not the protein transcription factors only. In essence, the following experiment will investigate the actual process of transcription by transferring a plasmid which contains an antibiotic resistant gene(Carninci, 32). If the deductions are correct, the plates that will contain ampicillin will demonstrate some degree of development, either colonial or growth, while that which does not will not exhibit any form of growth. The apparent lack of growth is attributed to the lack of resistance to the antibiotic that occurs due to lack of appropriate plasmids.

Methods

A vial which contained a solution of calcium chloride (aqueous) and a tube containing a strain ofE. Coli were both immersed in an ice bath. The aqueous Calcium Chloride was then transferred to a tube that contained 5µl of bacteria. The amount of Calcium Chloride Solution transferred was 590µl. The transfer was achieved using a sterile pipette. The solution was mixed using an index finger that tabbed the tube. After10 minutes of incubating, the cells became ‘competent’ since they could take in DNA from the provided medium. The cells were left stored in the Calcium Chloride solution for some time to allow for proper competency. In essence, Calcium Chloride is an essential reagent as per the experiment.

Two different tubes which were visually labeled ‘C’ and ‘lux’ were immersed in an ice water bath. Tube ‘C’ contained controlled plasmid plasma while the one labeled ‘lux’ was for the plasmid lux. A pipette, which was sterilized, was employed to add 5µl control plasmid to the tube labeled “C” and 5µl plasmid lux to the one which was labeled “lux.” Both of these tubes were then maintained within an ice bath. A transfer pipette (sterilized) was employed to add 70µlof the solution containing competent cells into each of the two tubes. It is important to note that each of the solutions created was tabbed with the index finger to ensure proper mixing of the contents. The two tubes were allowed to rest for about period of fifteen minutes, meanwhile, 35µl of the competent cells were dropped to a tube which was then labeled “NP” meaning that it lacked plasmid. After the fifteen-minute duration, heat shock was done by subjecting the tubes to a 37°C preheated water bath for another fifteen minutes. A pipette was employed to drop about 275µl nutrient broth to both the lux and the control tubes. An additional one hundred and fifty µl was put into the NP tube. The cells were then subjected to incubation at the optimum temperature. The solutions were then spread into different plates depending on the treatment, i.e.,LBNP, LBlux,LB/AmpC, LB/Amplux,LB/AmpNPand LBC. These 6 agars were incubated, and the results were determined after the specified period.

Results

The results were obtained from prior to the observation of the cultured media after incubation. Colonial growth represented an increased growth rate while lawn growth represented a relatively decreased one.

Process/

Growth Type Seen

Reasoning

LBc

Lawn

Ampicillin was absenthence nogrowth restriction

LB/Ampc

Colonial

Precence of Ampicillin restricted the growth of bacteria

LBNP

Lawn

Ampicillin was absent hence no growth restriction

LB/AmpNP

Colonial

Precence of Ampicillin restricted the growth of bacteria and the celllacked. Plasmid to make the bacteria antibiotic resistant

LB/Amplux

Colonial

Precence of Ampicillin restricted the growth of bacteriaLux is responsible for Bioluminescence

LBlux

Colonial

Ampicillin was absent hence no growth restriction, Lux responsible for Bioluminescence

The above table demonstrates the results obtained after the experiment. The results are as expected since no experimental errors were apparent. The LBC, LBNP as well as LBluxplateshad no ampicillin to restrict the growth of the bacteria. The type of growth observed is hence conclusively lawn. In addition to the above, no bioluminescence observed in LBC and LBNtreatments. The lux in the LBluxtube was responsible for the luminescence.

Discussion

The function of the calcium chloride is to neutralize the negative charge which exists on the cell wall for thesugar-phosphate group DNA group to be near it. This process was employed to make the cells competent. Prior to the above, there was a need of a way that the DNA can be made to enter the cell. This was possible through the process of heat shock. The overall process serves to give the bacterial entity some additional energy that will allow it to break open the pores within the cell so that the plasmid DNA can come through. The pores in question are those found along the margin of the cell wall. The plasmid that was allowed to pass through solely consisted of an ampicillin resistant gene and a lux operon.

The success of the transformation process can be identified after three conditions have been met. First, there is a cell that the specialized DNA can be immersed, there are a means of delivering the unique DNA into the existing host cell in question, and finally, there is a way of identifying transformation that would be apparent regarding the growth of the culture. In light of the experiment, all three conditions were met. If the bacteria were able to survive in an environment containing ampicillin, the bacteria had acquired a gene for ampicillin survival that, in turn, enabled it to survive in that situation. There are a variety of factors which influence the overall efficiency of the transformation process. These are apparently the number of bacteria and also the amount of plasmid DNA which have been exposed to each other. The degree of heat used during the experiment also influences the efficiency of the experiment to some extent. This is because less heat signifies an impermeability of the cell wall hence less genetic content can enter through.

In both LBNPand LBC agars, there were relatively an increased number colonies. This observation is due to the apparent absence of ampicillin that would in turn hinder their growth. In the LBc and LB/AmpNPagars there was lesser number of colonies compared to the first two; this is because LB/AmpNP contains ampicillin which restricted the growth of colonies. There were nonotedgrowths in LB/AmpNP and LB/AmpC agars due to the precence of ampicillin.The LB/AmpCand LBCagars exhibiteda lesser growth of colonies because LBC did not have ampicillin while the LB/AmpC had. A similar case can be said forLB/AmpluxandLBNP. There was no ampicillin in LBNP to restrict the growth of colonies, LB/Amplux had ampicillin thatrestricted the same. As a result, there will be few colonies. In the case of LB/AmpNP and LB/Amplux, there were no growths of colonies because both contain ampicillin that restricts the growth of colonies.

Works Cited

Bruijn. Stress and environmental regulation of gene expression and adaptation in bacteria. Hoboken, New Jersey: John Wiley & Sons, Inc, 2016. Print.

Bullerwell, Charles E. Organelle genetics : evolution of organelle genomes and gene expression. Berlin Heidelberg: Springer, 2012. Print.

Carninci, Piero. Cap-analysis gene expression (CAGE) : the science of decoding genes transcription. Singapore: Pan Stanford, 2010. Print.

Huang, Suming, Michael D. Litt, and C A. Blakey. Epigenetic gene expression and regulation. London, UK: Academic Press is an imprint of Elsevier, 20fifteen. Print.