Tryptophan Synthesis

The purpose of the promoter sequence is to initiate the expression of the transgene in the cells of the relevant tissue. The promoter sequence of the enviropig is obtained from the parotid secretory protein (PSP) gene sequence of a mouse, and it initiates the expression of the transgene, Escherichia coli’s phytase gene, in the secretory cells of the salivary glands of the pig. The transgene codes for the protein that is required for the expression of the gene of interest, in this case the Escherichia coli phytase gene code for the phytase protein. Finally, the mouse PSP terminator sequence is used to signal the end of the gene/code for the protein of phytase. Enviropig’s gene construct does not have an antibiotic resistance marker gene, and the purpose of this gene is to identify the cells with the gene construct.

Then the tRNA molecules link together and transfer the amino acid to the ribosome. An Anticodons pair with a codon takes the

E. coli is a single chromosome (haploid). E. coli has 4600 kilobases with about 4300 coding sequences and has around 1800 known E. coli proteins. 70% of its genes are monocistronic and 6% are polycistronic, roughly having the same amount of genes on each strand [2]. E. coli is a member of normal flora that inhabits humans and other animals. E. coli is heterotrophic and usually obtains its food from the organism it is living in. E. coli is dependent on carbon; its main source of carbon is obtained from glucose. E. coli is a facultative anaerobe meaning that it can use oxygen and other electron acceptors as the final acceptor in an anabolic or catabolic pathway. The optimum temperature of E. coli is 37o C,

1. EMG 9 and EMG 26 contain strain _lac-_(I- Z+ Y+) and strain _lac -_ (I+ Z- Y-)respectively.Three genes huddled together on the chromosome are required for two strains of _E.coli_ to utilize lactose.Consisting of three genes, namely, _lacZ_, _lacY_ and _lacA_, the _lac_ operon orderly handles these genes to code specific enzymes necessary for the metabolism of lactose. The genes _lacZ_, _lacY_ and _lacI_ would code for beta-galactosidase, galactosidase permease and _lac_ repressor respectively. Regulation of _lac_ operon is also tight and the operon's negative control is made possible by _lac_ repressor (Hill, 1996). With the presence of _lac_ repressor, _lac_ operon will be deactivated and will

In the pGLO Bacterial Transformation lab, Escherichia coli is transformed with a gene encoding green fluorescent protein by inserting a plasmid containing the GFP gene, beta-lactamase, and arabinose into the bacterium. Successfully transformed bacteria will grow in the presence of ampicillin and glow a bright green color under ultraviolet light. The sugar arabinose is responsible for switching on the GFP gene in the transformed cells, without it, the gene will not be expressed.

For this experiment, E. coli was best for genetic engineering because of their size, and their fast reproduction (Spilios, 2017). E. coli will be genetically transformed using an engineered plasmid. A plasmid is a circular piece of DNA which independently replicates and multiplies because it has its own origin of replication (Spilios, 2017). The pGLO is the plasmid used in this experiment. Plasmids are used as vectors and they contain manipulated genes such as genes coding for antibiotic resistance for drugs like ampicillin. This antibiotic resistance of such serves as the selectable marker in genetic transformation and for genetic transformation to proceed, the cell must reach competency which is the physiological state that is required for the vector plasmid to get into the cell for transformation (Spilios, 2017). While competency can be reached naturally in some organism, it must be reached artificially in E. coli through treatment with CaCl2 and exposing them to heat shock using incubation (Spilios, 2017).

Escherichia coli, or E. coli, is a common bacterium that can be found in diverse environments all over the planet, including the gastrointestinal tracts of animals and humans. Many of these strains of E. Coli are essential mechanisms in the digestive tract, while others are pathogens that can cause complications in urinary and intestinal tracts. (Payne & Sparks) In research, E. Coli is commonly used as a model organism, meaning they are widely studied by scientists for a variety of purposes due to their experimental advantages. E. Coli is comparatively simple, and there are many advantages to studying these prokaryotic cells in the fields of biochemistry and molecular biology. E. Coli has this simplicity and is relatively easy to propagate in a lab environment. Their genome has been completely sequenced and many things we know about DNA, protein synthesis, and gene linkage have been derived from studies regarding this particular organism. (Cooper)

The gene for GFP is regulated by an inducible repressor system, meaning it is turned on by environmental factors. In this case, GFP is expressed when exposed to the sugar arabinose; moreover, the organism will glow only when arabinose is present. Without arabinose, the repressor attaches to the regulator/ operator segment of the gene and acts as a roadblock, preventing RNA polymerase (RNAP) from reading the structural gene and creating a complementary mRNA strand; thus, GFP protein production is prohibited. However, when present, arabinose binds to the repressor, changing its active site, so it can no longer bind to the operon; RNAP can then initiate the transcription of the GFP gene by binding to the promoter segment. Thus, I predict that the E. coli will glow only when pGLO and arabinose are present because arabinose activates the expression of GFP, a gene found in pGLO. Regulation of gene expression is crucial to prevent wasteful overproduction of unneeded proteins and to allow adaptation to new environments (Regulation of

Bacteria belongs to a group of organism that lacks cell nucleus and membrane bound organells. This group of organisms are termed as prokaryotes. Prokaryotes follows the central dogma of molecular biology first proposed by Francis Crick in 1958 to synthesize proteins from mRNA through a process called translation and the mRNA is being synthesized from the DNA by another process called Transcription. Temperature, nutrient availibity are some key factors that start the process of synthesizing proteins in response to these key factors. Example. This paper will provide an explanation as to how bacteria decode the genetic information to produce

Prokaryotic genes are often polycistronic which means that each gene has a start site so each is transcribed independently. However, directly neighboring genes are transcribed in conjunction. Operons are a collection of contiguous genes that code for metabolic enzymes whose transcription is controlled by one operator (Jacob & Monod, 1961). Since operons are coded in one direction, the upstream genes can inhibit the transcription of later genes (Hartl, 2011).

The instructions tell the cell what to do and what traits to display. In this particular lab, pGLO plasmids was inserted into E.Coli bacterial colonies. This plasmid contained the bla gene, the araC gene and the GFP gene. These genes changed the bacteria by allowing it to be resistant to a particular antibiotic, Ampicillin and also to glow fluorescent green when in the presence of arabinose and when shown under a UV light. The bla gene codes for the beta lactamase enzyme which allows the bacteria to be resistant to Ampicillin, while the GFP gene codes for the protein that glows in the presence of UV light. Also, the araC gene codes for the protein that allows the GFP gene to produce the green glow protein only in the presence of arabinose. When the bacteria was in the presence of ampicillin and arabinose, the arabinose acted as an inducer which led the GFP gene to start producing GFP. This led to the bacteria glowing under UV light. This clearly shows gene expression as the GFP produced a protein after it was induced. I also learned from this lab that a cell’s DNA can be changed in order for it to produce different proteins. I also learned more about inducers, which bind to repressors to allow a gene to produce a protein. Furthermore, this lab also reinforced my knowledge of of how protein synthesis can be stopped or activated. Overall, this lab shows gene expression and is a primary example of how a cell’s traits can be

The experiment was conducted in two main parts: in the first part, the single and double restriction digests were run to cut the plasmids using combinations of three different enzymes. In the second part, the fragments resulting from each reaction were separated using gel electrophoresis. 1) Restriction Digests In this part of the experiment, seven different samples were prepared with different combination of reactions. Given the unknown plasmid and three restriction enzyme (HindIII, BglI and PvuII), seven different Eppendorf tubes were prepared to account for 3 single digests, one with each of the three restriction enzymes, 3 double digests combining two enzymes at a time and alternating between all the possible combinations, and also for the control reaction where no enzymes were added to the plasmid.

Organisms cannot depend solely on spontaneous reactions for the production of materials because they occur slowly and are not responsive to the organism's needs (Martineau, Dean, et al, Laboratory Manual, 43). In order to speed up the reaction process, cells use enzymes as biological catalysts. Enzymes are able to speed up the reaction through lowering activation energy. Additionally, enzymes facilitate reactions without being consumed (manual,43). Each enzyme acts on a specific molecule or set of molecules referred to as the enzyme's substrate and the results of this reaction are called products (manual 43). As a result, enzymes promote a reaction so that substrates are converted into products on a faster pace (manual 43). Most enzymes are proteins whose structure is determined by its sequence of its amino acids. Enzymes are designed to function the best under physiological conditions of PH and temperature. Any change of these variables that change the conformation of the enzyme will destroy or enhance enzyme activity(manual, 43).

Introduction The purpose of this experiment was to chemically develop Nylon-6,6 by combining adipoyl chloride to the 1,6-hexanediamine solution. According to the American Chemical Society, polymers are created from repeated pieces that are connected together.1 The pieces are called monomers. The synthesis of polymers is called polymerization.

The regulation of transcription of most aminoacyl tRNA synthetase genes in Gram-positive bacteria is controlled by a common transcription anti-termination system (cite?). This anti-termination system is formed by the T-box riboswitch, which is characterized by a conserved RNA sequence and secondary structure elements. T-box riboswitches are located in nascent 5 ' RNA transcripts (leader RNA) of these aminoacyl tRNA synthetase genes. Synthesis of the full-length mRNA requires stabilizaton of the anti-terminator motif. This is facilitated by RNA-RNA interactions between cognate uncharged amino-acyl tRNAs and leader RNA (Grundy et. al, 1994). Specifically, in the case of glycine tRNA synthetase, the specifier sequence (GCC) of the leader RNA base-pairs with the tRNA anticodon (CGG), and the antiterminator element pairs with residues in the 3 ' acceptor end of the tRNA. Interestingly, only cognate uncharged tRNAs promote antitermination (Grundy et. al, 1994). For example, the tyr-tRNA synthetase gene is only transcribed when uncharged tyr-tRNA 's are present (Grundy and Henkin, 1993). Thus, transcription is induced in response to high levels of uncharged tRNA 's due to a deficiency of the corresponding amino acid or aminoacyl-tRNA synthetase. This logical regulation of transcription thereby tries to restore levels of charged amino-acyl tRNA synthetases. The exact requirements for these tRNA-leader RNA interactions that induce transcription have not yet been fully defined,