More Than Meets The Eye; Identifying A Diseased Gene With PCR

A) DNA testing is when one uses science to discover more about themselves than is possible by looking at oneself. Such a test can decode one's entire genome and can thus indicates one's genetic future. Genetic tests are done using Polymerase Chain Reaction (PCR), a process which takes small DNA sequences, uses a DNA polymerase, a primer, and nucleotides to accomplish its goal. DNA tests are done for a variety of reasons, including: identifying other species, and discovering one's genetic future. PCR can be used to sequence a gene of interest and allow scientists to determine whether the gene is diseased (harmful) or not.

B) The purpose of this experiment is to determine whether each of us possess a "diseased" gene. The gene is not a real diseased gene, but it will offer insight into our evolutionary past and give us real world experience in the realm of genetic testing, thus teaching us the significance of such testing.

C) To complete this experiment, our lab table will need to complete a number of tasks correctly. We will need to extract our DNA through cheek cell extrcation, which requires saline mouthwash for extraction. Upon extracting the cheek cells, they will need to be broken open which we will utilize a lysis buffer for. After breaking open the cells instagene matrix will be added to kill DNASE. Next we will utilize PCR (Polymerase Chain Reaction) to amplify the desired sequence of DNA. After PCR, we will use Gel electrophoresis. The Gel electrophoresis will allow us to determine whether we are each individually carriers of this "disease" by visually comparing the Gel and the lanes within it to one another.

D) The objective of this experiment is to figure out whether we are the carriers of this "diseased" gene. I am unable to produce an accurate hypothesis at this moment, but if I was to attempt to do so, I would say that I am not afflicted with or a carrier of the "diseased" gene.

Genetically Modified Organism Discussion

Results: 

Our gel did not reveal the results of this test, so our experiment did not work. However, we were able to analyze the gel of fourth period table two, and the results indicated that their test food was not genetically modified. Their number three band corresponded directly with their number five band on the gel, thus indicating its organic status. Even though the test food appeared to be organic, Mr. Chugh's corn flower band (number four) corresponded with the number six band on the gel. This means that The cornflower (test food two) is genetically modified.

 

Sources of Error:

-Ran Gel for too long, so DNA and the loading dye ran off the Gel.

-Did not crush test food well enough, causing the chunks to be too big to fit into the pipette, which caused no lettuce to be analyzed by PCR or Gel Electrophoresis.

-Accidentally extracted leftover DNA from the previous class's mortar and pestle.

-Added too little loading dye, and the DNA is not visible on the gel.

-Missed the wells when adding the DNA into the gel.

Oh No GMO; Testing Our Favorite Foods To Inspect Prevalence Of GMOs In Agriculture

A) GMOs are genetically modified organisms, and their use in agriculture is increasing, and becoming harder to distinguish. Genetically modifying plants can make them more durable, as some of the traits they can gain from genetic modification include resistance to: frost, drought, disease, and insects. GMOs are made by placing a gene of interest into a tumor inducing plasmid, adding the plasmid into agro-bacteria through the method of heat shock, and infecting a plant cell with the bacteria. The plant cell will then divide and grow up into a large, healthy plant. Genetically modified organisms can be identified by taking a plant cell, and finding a specific gene sequence located in a tumor inducing plasmid inside of the plant cell. This process is made possible by Polymerase Chain Reaction (PCR), a process which takes small DNA sequences, uses a DNA polymerase, a primer, and nucleotides to accomplish its goal. The controversy behind GMOs is the possibility that they could in fact harm the public. One way is that by making plants stronger, scientists are in fact strengthening the bugs and weeds which feed on plants. For example, weeds that crossbreed with herbicide resistant crops could evolve into super-weeds, and bugs which feed on toxins in plants will no longer be susceptible to such poisons and evolve into super-bugs. Another way is the possibility of allergies or religious conflict that could arise from people unknowingly consuming plants containing proteins from an animal which is not revealed (ex. somebody allergic to fish eating frost resistant tomatoes and becoming deathly sick from the fish protein). GMOs can be a source for good, and could end world hunger and famine, yet the risks and potential dangers must be weighed before any such actions can be taken.

B) In this lab, we will be testing our various favorite foods to identify whether or not they are genetically modified. While doing this we will be policing our own food, and deciding for ourselves whether we really want to eat food which has been altered genetically. This can be applied to the population of the world, and could allow people to make this decision even if the foods are unlabelled.

C) To extract the DNA, we will be heating a hot water bath to 99 degrees Celsius. This should effectively burst the cell membrane. DNASE is a protein developed by Eukaryotic cells to kill DNA outside of the nucleus. To get around this problem, we will add Instagene. Instagene matrix beads are designed to kill DNASE, thus allowing us to perform the experiment. We will use a sequence of interest (from DNA) from the Tumor Inducing plasmid (one that would not occur naturally in the plant) through PCR, to identify if genetic modification has taken place. Special techniques that we will be using include the use of Polymerase Chain Reaction. Our controls of this experiment are the Non-GMO food control with plant primers, Non-GMO food control with GMO primers, the GMO positive control with plant primers, and the GMO positive control DNA with GMO primers. They serve as a comparison for the unknown pieces of vegetable or fruit, and also as an indicator of whether the experiment worked successfully or not. We will be using Gel Electrophoresis in this lab to visually compare the similarities and differences of GMO/Non-GMO foods with the foods that we brought in to experiment with. Depending on the food, we should have accurate results of how much food is genetically modified in our supermarket.

D) In this experiment, the main objective is to see how much of the food is genetically modified. So, the variables of this experiment will be the food we bring in to test (Test food with plant primers, and Test food with GMO Primers). My hypothesis for this experiment is that  both plants which we test will be genetically modified. I believe this because, 70% of the food in the united states is genetically modified, and odds are that both will contain some type of modification.

How To Make Your pGLO; Transformation Of Fluorescent Sea-jelly Genes Into Bacteria

A) Transformation is a genetic process which means change caused by genes, and involves the insertion of foreign DNA into an organism to change its trait. Transformation is carried out by restriction enzymes to cut plasmid DNA and insert new code in its place, and heat shock (rapidly heating and cooling cells with calcium chloride) which will move the plasmid (DNA which contains one or more genes which would be beneficial to a bacteria's survival) into the cell. It also is a key instrument of biotechnology, and is used in numerous ways, such as, bioremediation (the genetic manipulation of bacteria to digest oil from spills), and medicine (use of gene therapy to correct defective genes). Also, another use of Transformation is to make bacteria glow by inserting the Green Fluorescent Protein (from Seajellies) into bacterial cells.

B) The purpose of this experiment is to insert the GFP gene into a plasmid, and move the plasmid into bacterial cells. This will cause the cells to produce Green Fluorescent Protein, and they will glow brilliantly. We hope to have a flourishing fluorescent colony by the end of this experiment.

C) In this lab, we will have four petri dishes: +pGLO and LB/amp, -pGLO and LB/amp, -pGLO, and LB, and the +pGLO and LB/amp/ara. We will use Transformation, which is carried out by restriction enzymes to cut plasmid DNA and insert new code (GFP gene) in its place, and heat shock (rapidly heating and cooling cells with calcium chloride) which will move the plasmid (DNA which contains one or more genes which would be beneficial to a bacteria's survival) into the cell. The controls of this experiment are the -pGLO, and LB petri dishand the -pGLO and LB/amp. The variables will be +pGLO and LB/amp,and the +pGLO and LB/amp/ara. I predict that the +pGLO and LB/amp will be the only bacterial colony to exhibit fluorescent characteristics. This is because it contains the ampicillin resistance gene, and the plasmid containing the GFP gene.

Discussing The Strategic Use Of Microarrays In The Study Of Lung Cancer Genes

A) Results: From the gene chip our lab group was able to determine that: Genes 1, 2, and 5 were active in in the cancer cells, Genes 2,3, and 6 were expressed in cells obtained from healthy tissue, and also that Gene 4 was express in neither the healthy cells nor the cancerous cells. We were able to tell this becauseof the colors of each gene on the slide. Genes 1 and 5 were pink (pink means expressed only in cancer), Genes 3 and 6 were blue (blue means only expressed in healthy cells) , Gene 2 was purple (this represents expression in both healthy and cancerous), and Gene 4 was clear (expressed in neither).

B) Sources of Error:
- Not enough of the cDNA was added, and the true color of the gene was not shown.
- The genes from the gene chip were not accurate, and we tested the expression of the wrong gene.
- We pipetted too much of the cDNA, and the slide overflowed, thus we were unable to get an accurate reading of the expression of the certain gene.
- We left the gel in the pipette for too long so it hardened and less than 20 microliters was exposed to the slide.

Translating Diseases; The Strategic Use Of Microarrays In The Study Of Lung Cancer Genes

A) Microarrays have simplified how scientists study gene expression in thousands of cells. This new technology detects patterns in transcription and translation, and aids in the understanding of the normal and abnormal aspects of cell function. The process of using Microarrays to analyze/study genes involving lung cancer include: creating a DNA chip, and using the DNA chip to analyze complementary DNAs isolated from both cancerous and noncancerous tissue (the same tissue). The Microarray will then indicate the differences between the two corresponding genes. This will be an indication of the extent to which transcription occurred. Some scientists have indeed observed that some genes are not transcribed as much in cancer cells as in healthy cells, and that some genes are not transcribed as much in healthy cells as in cancer cells.

B) The purpose of this experiment is for our lab table to use Microarrays to study Genes Involved in Lung Cancer. This will hopefully give our group a better understanding of the causes of the disease on a genetic level, and thus allow us to speculate with possible methods to prevent or cure it.

C) In this experiment, our lab table will be studying six genes using a Microarray. We will prepare the Microarray by spotting six different gene sequences onto a glass slide. Then we will obtain DNA tubes from a 70 degree water bath, and spot the appropriate gene solution onto the correct areas of out slide. To hybridize the Microarray, our group will add fifteen microliters of hybridization solution to each spot. We will then wait and observe our results when they appear.

D) Although we have the six genes that we are to study, making predictions is not possible because we do not have the background knowledge to hypothesize our results. However, I am quite certain that our group will accomplish something worthwhile, and we will learn valuable facts about gene expression and the effects of cancer on this.

The Sticky Ends Justify The Means Discussion

A) During Day one of the lab, our lab table acquired the DNA of the five suspects, and the DNA found at the crime scene. The purpose of the experiment was to identify the culprit by matching the DNA found at the scene with the DNA of the five suspects using Restriction Fragment Length Polymorphism. After acquiring the DNA, we added restriction enzymes to the tubes, flicked/tapped them, and placed them in a hot water bath for 30 minutes. On Day two we ran the newly cut DNA through a gel, and visually compared the samples using loading dye to stain the DNA (make it visible), and attracting the fragments to the positive end through Gel electrophoresis. Gel electrophoresis functions by pulling fragments of DNA (negatively charged) through the gel matrix towards the red end (positive), run to red. On Day three, our group was able to compare the DNA of the five suspects with the DNA found at the crime scene. We were able to positively identify Chloe Krey as the culprit in this crime scene.

B) Possible Sources of Error:
- We did not add enough loading Dye so the DNA fragments were not visual.
- Accidentally scraped off a skin cell into a test tube, and my DNA was analyzed instead of a suspect's DNA.
- We did not add enough DNA to the tube, and thus not enough substance was present for DNA to become fully visible.
- Accidentally added the wrong suspects DNA into the wrong well, and the wrong suspect was 'proven' guilty by the DNA evidence.
- Accidentally added the food coloring in place of the Loading Dye, and the DNA was not properly stained thus leading to an inaccurate reading.