Human Mitochondrial

The mitochondrial (mt) genome which contains only 37 genes, is used for the process of the production of energy and its storage in ATP. In the "Human Mitochondrial DNA Kit" background, it said that there was strong evidence that mitchondria once existed as a free living bacteria, which were taken up by primitive ancestors of eukaryotic cells. The MT genome has some bacteria like feature, is also a circular molecule and very few noncoding sequences (introns) interrupt mt gene. In 1981, the entire DNA sequence of the mt genome was determined. In the mt genome, there is a region of noncoding nucleotides (1,200) in which it contains signals that control replication of the chromosome and the transcription. So this DNA sequence is called the hypervariable because it accumulates mutations at approximately 10 times the rate of nuclear DNA, and because of this, ther is a unique pattern of single nucleotide polymorphyism, which could be inherited through generations. Also in the 1980's, Alan Wilson of UC Berkeley, used mtDNA polymorphisms to create a family tree, that showed the ancestral relationships of modern people. From this they were able to conclude that all modern humans arose from Africa about 200,000 years ago. Today mt DNA has been used to identify the remains of unkown soldiers in Vietnam, also determine the reamins of the Romanov royal family and the relationship of Neandertal remains to modern humans
Directions:
Step 1. First we labeled one screw cap tube containing 200 hl of InstaGene matrix and then rinsed our mouth with saline solution for 30 secounds. (chewed inside of our gums to loosen cheek cells.).
Step 2. Then we placed this saline solution that we spit out into the cap tube containing the InstaGene and then we centrifuged it for about 2 minutes.(After it was centrifuged, there was a white bead of whitish cells at the bottom of the tube.)
Step 3. Then after centrifuging, we poured out the extra saline solution out of the tube, so then we only had the pellet of white cells at the bottom of the tube.
Step 4. Then we vortexed the tube so there would be no clumps of cells remaining and to resuspend the pellet of cells.
Step 5. Then we transfered the resuspended cells, into the screwcap tube containing InstaGene.
Step 6. Then we vortexed the tube and then placed the tube into a water bath (56 degrees Celcius) for 10 minutes.
Step 7. After we placed the tube in the vortex again and then put it into a boiling water bath (100 degrees Celcius) for 5 minutes.
Step 8. Then the next day we placed the mixture into a PCR tube and also added some yellow master mix. Then we placed it into the thermal cycler for 40 cycles.
Step 8. Finally, we added loading dye to our PCR tubes and then placed our mixtures into the wells of the agrose gel and then turned on the electrophoresis apparatus for about 10 minutes.
Results: Our results showed that we were all negative for the disease, but it could possibly be wrong because we could have messed up when loading the mixtures into the wells of the agrose gel. Since our results were very faint and very hard to see if they really were negative for the disease. I also think that my tube might have not had enough cheel cells because when I tried transfering the cheek cell bead to another tube, it sort of feel apart.So this may have caused me to transfer more of the saline solution rather than my cells.

DNA Testing

Background: In todays modern world, we use Molecular biology, is the study of of genes and other molecular structures, that regulate the flow of genetic information from generation to generation. Also Biotechnology uses this information to help solve human problems, by taking this information and testing it through DNA tests, just like our very own lab. In our our lab we are doing a DNA test, to find out whether or not certian people in class have a specific gene that we are tryign to target. This DNA testing is used for all sorts of things, but it is most commonly used for helping people find out if they have or carry a certain disease like Huntington disease. In order to do this lab we must use PCR, which is a technique that takes a trace of DNA and produces a large amount of it. This technique today is widely used in biotechnology and it even helepd transfrom molecular biology into a multidisciplinary research field within five years of it being made. PCR has also had a profound impact on four main areas of genetic research: gene mapping, gene cloning, DNA sequencing and gene detection.
Directions:
Step 1. First we labeled one screw cap tube containing 200 hl of InstaGene matrix and then rinsed our mouth with saline solution for 30 secounds. (chewed inside of our gums to loosen cheek cells.).
Step 2. Then we placed this saline solution that we spit out into the cap tube containing the InstaGene and then we centrifuged it for about 2 minutes.(After it was centrifuged, there was a white bead of whitish cells at the bottom of the tube.)
Step 3. Then after centrifuging, we poured out the extra saline solution out of the tube, so then we only had the pellet of white cells at the bottom of the tube.
Step 4. Then we vortexed the tube so there would be no clumps of cells remaining and to resuspend the pellet of cells.
Step 5. Then we transfered the resuspended cells, into the screwcap tube containing InstaGene.
Step 6. Then we vortexed the tube and then placed the tube into a water bath (56 degrees Celcius) for 10 minutes.
Step 7. After we placed the tube in the vortex again and then put it into a boiling water bath (100 degrees Celcius) for 5 minutes.
Step 8. Then the next day we placed the mixture into a PCR tube and also added some yellow master mix. Then we placed it into the thermal cycler for 40 cycles.
Step 8. Finally, we added loading dye to our PCR tubes and then placed our mixtures into the wells of the agrose gel and then turned on the electrophoresis apparatus for about 10 minutes.
Results: Our results showed that we were all negative for the disease, but it could possibly be wrong because we could have messed up when loading the mixtures into the wells of the agrose gel. Since our results were very faint and very hard to see if they really were negative for the disease. I also think that my tube might have not had enough cheel cells because when I tried transfering the cheek cell bead to another tube, it sort of feel apart.So this may have caused me to transfer more of the saline solution rather than my cells.

Genetically Modified Plants

      Genetically modified products today, are widely used in the U.S., in order to produce a better food product that can withstand harsh environments and other factors such as insects. Proponents of GMOs, believe that these plants hold the solution to to help put a stop to overusing pesticides and herbicides, that are causing grave health conditions to us and problems to our environment. Farmers have been genetically modifying their crops for centuries through crop breeding, in order to encourage specific traits, like allowing a plant to bear more fruit. But today, there is the option of selecting specific genes from anything, not just from another plant, but also from organisms. Because of this new technology farmers today, are able to produce plants that have delayed fruit ripening , resistance against fungi and even increase crop yield. Though many people today, believe that GM plants are bad because the plants are created naturally through evolution and many believe it could cause health problems to humans and create super resistant bugs and weeds. Today there are two methods of identifying if a crop is a GM plant, one method is using an enzyme called immunosorbent assay, which identifies proteins. So this enzyme, basically identifies specific proteins produced by a GM plant (can only be used on fresh produce.). Another way to test for a GM plant, is using polymerase chain reaction (PCR), which identifies sequences of DNA that have been inserted into a GM plant. In our lab, we are going to test our two foods that we brought in from home, to see if it is a GMO food/plant.

 Procedure:

Day One: First we weighed out .5-2 g of certified non-GMO food and our food that we brought in, and put it into a mortar. After, we added 5 ml of distilled water to every gram of food and then grinded it with a pestle for two minutes. Then we pipeted 50 ul of the mixture into a screwcap tube containing 500 ul of InstaGene. After this we placed our test food tubes into a 95 degree Celcius water bath for 5 minutes, then placed the tubes into a refrigerator, until the next day.

Day Two: On day two we numbered our PCR tubes 1-6, corresponding to the tubes content. The even numbered tubes got the red master mix (20 ul of master mix to each tube.), and the odd numbered tubes got the green master mix. Then we placed the tubes into an ice bath and put in master mix according to each tube, and then we placed them into a thermal cycler.

Day Three: On day three we used electrophoresis (which we placed each test tube mixture into a specific well), and then after this we were able to determine if our test foods we brought in were GMOs.



Results: I predicted that our test food, which was an orange, wouldn't be a GMO because it was grown in one of our backyards, but we found out that it was actually a GMO.  Our results showed that both of our test foods tested positive as a GMO product. Our lab could have possibly flawed though because when we were inserting our mixtures into the wells in the agrose gel, one of the mixtures in a well sort of spilled out alittle.

pGLO Lab

In this lab we are using the technique called genetic transformation, in order for us to create a bacteria that glows using a gene that codes for green fluorescent protein. Genetic transformation is used, by inserting  a gene of interest from another organism into another, in order to change an organisms genetic makeup. Today, genetic transformation is used in biotechnology in many ways, some uses of this technique are to create perfect fruits and vegetables. Today in our lab, we are using a green flourecent protien that comes from jelly fish in the dark waters of the ocean, and we are going to take this gene and insert it into a plasmid and then insert it into our bacteria. (A plasmid is a circular DNA molecule capable of self replicating itself and also able to carry genes.)