#1. a) The Use of a Bacterial Plasmid to Clone and Sequence a Human Gene
The process begins with restriction endonucleases scanning and binding to double-stranded DNA at specific base-pair sequences, the recognition sites, in a predictable manner. The restriction sites are usually 4 to 8 base pairs long and are characterized by the palindromic sequences, with both strands having the same sequence when read in opposite direction. After the restriction endonuclease binds, it starts to disrupt, using hydrolysis, the phosphodiester bonds between neighbor nucleotides, causing the H-bonds between base pairs in the cutting region to be broken. This cuts the original double-stranded DNA strand, producing two DNA fragments, which may differ for different restriction endonucleases, depending on where the phosphodiester bond is broken when cut by the endonuclease. This process can produce either blunt ends (where ends of the DNA fragment are fully paired with no overhangs), or sticky ends (where both DNA fragments have nucleotides lacking complementary bases and overhangs are produced). However, sticky ends are more useful for genetic engineering. The next step, gel electrophoresis, separates the gene that has been excised, from the unwanted fragments taking advantage of chemical and physical properties of DNA. The DNA fragments travel through gel as a result of charge passed through it causing the longer fragments to separate from shorter ones, which helps in identifying gene and makes it easier to cut it out from the gel. The DNA fragment with the desired gene is, therefore, excised and purified. The same restriction endonuclease, that is used to cut the original DNA strand, then splices this gene into a plasmid (small, circular DNA molecules found in bacteria). Because the plasmid and the foreign gene are cut by the same restriction endonuclease, the sticky ends formed, are complementary and anneal to each other forming H-bonds. The DNA ligase reforms the phosphodiester bonds, after which, the recombinant plasmid with the foreign DNA, is introduced into the bacterial cell, in the process of transformation, and replicates to form clones (exact copies of itself). Overall, this technique of altering sequence of DNA molecules became very useful for many reasons, one of which is production of hormones. Hormones such as insulin and somatropin, were produced by inserting a gene into a plasmid, and became essential hormones in medical practice. In this process, the needed human genes were incorporated into plasmids and activated or inactivated when needed, using specific inducers for promoter regions.
Figure 2 shows the results of the electrophoresis. Lanes 5 and 7 indicate the fragments obtained when the plasmids are digested with both restriction enzymes, indicating the approximate fragment size for the hlyA gene, the pK184 plasmid and the pBluescript plasmid. This is useful for identifying the recombinant DNA needed for this experiment
A recombinant plasmid are created by first using an enzyme that can identify and isolate specifically which gene that need to be cut. They are call restriction enzymes or restriction endonucleases, and more than 100 of these enzymes have been isolated. After the human gene (gene of interest) that codes for the desire trait is located on the chromosome restriction enzyme does it job, by cutting out the gene from the DNA. Now, the two ends of the human gene will be those that will link up with the open ends of the plasmid. An enzyme, DNA ligase, is used to couple each end of the gene to the open ends of the plasmid; this thus restores the circular DNA molecule with the human gene. Now the plasmid, with the human gene, is reinserted into the bacteria. They are then cultured and produced in large quantities of identical bacteria carrying the human gene. Now, these bacteria produce the human protein coded for by the spliced human gene. The protein is then isolated and purified and are ready to be injected into patients (crop, etc.) (Gish 1998).
The two modes of analysis that will be used to identify an unknown insert piece of DNA would be plating the transformation cells onto LA plates that have either ampicillin or chloramphenicol and PCR. We will use the PCR thermocycler to denature the restriction enzymes that were specifically used to assimilate the vector DNA. It is important to use the PCR thermocycler because denaturation of the restriction enzyme will prevent the restriction enzyme from cutting the vector DNA, after the insert DNA has assimilated to the vector DNA. After the addition of specific primers that complement the base pair to its corresponding target strand, PCR will be used. Subsequently, Taq polymerase will be used to determine whether the insert DNA has been properly assimilated to the vector DNA. Within this specific situation, the target strand will be the insert DNA. After we let the PCR thermocycler run for approximately 2 ½ hours, we will then put our PCR products in the gel and run the gel to completion. After the gel has run to completion, we will then take a photograph of the gel using the UV transilluminator with the assistance of our TA. If the insert DNA was properly assimilated to the vector DNA, then our corresponding gel photo would have one band. After the cells have been transformed, we would g...
A human DNA, in which biologists have identified and isolated the gene of interest using probes or antibodies, will then be chosen. This gene of interest is incorporated into the plasmid cuts. These new plasmids are mixed with, and taken up by bacterial cells under suitable conditions. As these bacterial cells reproduce, the plasmids containing the gene of interest will be copied, and transferred to the bacterial progenies. Genes are segments of chromosomes that code for specific polypeptide or RNA molecules. Plasmids are small loops of DNA separated from bacterial chromosomes, or viral vectors. Restriction enzymes are enzymes that cut DNA at highly specific areas that always contains the same sequence of
techniques in their laboratory to make copies of cells or organisms with a valuable trait.
I want to go to college for the wholesome intellectual stimulation and to get more involved with my community. College will be my last opportunity to do so before I enter the work force. At college I can be intellectually and academically challenged; I will be able to learn with friends who are as dedicated to learning and the pursuit of knowledge as I am. My goals in college are to acquire the skills that I need to lead a successful and productive life, to expand my knowledge of the world, to step out of my comfort zone, and to become more proactive.
Modern techniques , rather than the gene map , maps the map of the DNA within the gene itself : the positions of short sequences " marker " are used as markers signaling over the cromosssomas . Once a gene is discovered, it is necessary to unravel its base sequence prior to its function being studied . The sequencing has become easier with the development of methods for cloning the DNA - producing large amounts of identical fragments. In the method most widely used DNA sequencing , the chain is denatured into single strands . These are then used as templates for DNA synthesis , but such that replication to as the double helix reaches a certain growth in the mold base . In addition to provide DNA polymerase and the four bases, A - G -C- T, also using small amounts of these dideoxynucleotide bases. This is incorporated , as the normal bases, the double helix growth but prevent the continuation of the chain. The fragments are then separated by gel electrophoresis and the base seq...
Are genetically modified foods safe? Genetically modified foods are crop plants created for human or animal consumption using molecular biological techniques. These plants have been modified to enhance certain traits like increased resistance to herbicides or improve nutritional content. This process traditionally has been done through breeding, but is not very accurate. Scientists have been using biotechnology to implant the gene that makes the plants act the way they want them to. Genetically modified foods have advantages and disadvantages on the environment and advantages and disadvantages on society. We have to weigh the positives and negatives to see if genetically modified foods are healthy for us and if we really need them.
Noted authors, Brandon Chambers, is quoted saying, “If you are going to fear anything fear success. Think about what you are doing and when you succeed what life you will have.” There are several different reasons why I could stay home, work and not go to college; I could go to work every day and make more money for the house, it’s easier, and I would be less stressed. Now, on the other hand there are many reasons why I should go to school; such as further my education, make my family proud, and make myself proud. I am attending college for several different reasons. One reason is to further my education. I hate feeling like I don’t know something, I like being the person everyone comes to for information. Also, because I want to be better prepared for my major, I want to be better then the next person with the job credentials. I want to major in Social Work and Criminology. I dream to be a counselor or an clinical service social worker. I want to help people who need someone there for them someone who can guide them or even just talk too. Another reason is because I would be the first in I would be the first in my family to go to college. I feel in some type of way I am setting an example for my parents and my brother that they too can go back to school. Lastly is simply because I love school. I love knowing more and more each day. College is not for everyone, but I will be successful by grasping the benefits, preparing for the problems, taking heed from experts, and working on strategies to be successful.
My purpose and goals in attending college is to make something of my life. It is a good thing. No one wants to be a nobody. But sometimes it takes time in order for one to realize this. & nbsp; Upon graduation from high school, I, like the majority of others high school graduates, had no clue what direction my life was heading or even what I wanted to become.
In a laboratory scientist will use a process called gel electrophoresis to separate DNA fragments. The DNA is cut into different sized fragments as a result from using restriction enzymes. The different sized DNA fragments are organized injected on agarose gel with an added substance that helps it glow after the test. DNA is negatively charged. Electricity is producing a positively charged are and a negatively charged area. Opposites attract and as a result the negatively charged DNA will move quickly to the positively charged area. Smaller DNA fragments will run faster the larger DNA fragments. After the electricity is turned off smaller DNA fragments will be closer to the positively charged area and the larger DNA fragments will be farther from the positively charged area. While it is glowing scientist can take a picture of the data and record the results and compare DNA samples to look for any abnormalities.
2). As a result, this scientific experiment changed the relationship of humankind and nature by foreseeing the modification of DNA of bacteria, yeast, plants, and animals to discover new medicines and to provide solutions for inherited diseases (Le Vine, 1999, p. 2).
Growing up in a household where a mother did not finish high school and a father did not finish elementary is not an ideal up brining for any child. I grew up emphatically impoverished for the majority of my life and my parents tell me they attribute that to the life choices they made in their youth. Today I use these circumstances not to debilitate my confidence, but to motivate me to strive for what I once imagined insurmountable.
Seidel, Jr., George E. "Cloning." World Book Student. World Book, 2014. Web. 13 Feb. 2014. source 19
Secondly the gene has to be cut from its DNA chain. Controlling this process are many restriction endonucleases (restriction enzymes). Each of these enzymes cut DNA at a different base sequence called a recognition sequence. The recognition sequence is 6 base pairs long. The restriction enzymes PstI cuts DNA horizontally and vertically to produce sticky ends.