Biological Molecules
Experiment-Specific Questions
Experiment 1: Monosaccharide Test
1. Fill in the table below with the results from the monosaccharide test experiment, and your conclusions based on those results.
Results Monosaccharide Test
Solution
Initial Color
Color with Benedict's Solution
Color After Heating
Monosaccharide?
glucose solution
clear
blue
orange
yes water clear
blue
blue
no sucrose solution
clear
blue
blue
no fructose solution clear
blue
orange
yes
1. Benedict’s solution is added to white grape juice and heated. The color changes from blue to orange. Based on this result, what biological molecules are
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DNA General Concepts
1. How does gel electrophoresis sort DNA fragments? Electrical charges (positive to negative and so on). Also by size helps sort fragments as well. Small dna moves faster than larger ones do.
1. If each individual has such a small amount of DNA, how do the bands on the gel contain enough DNA to be visible? By restriction enzymes then amplified by polymerase chain reaction to make many to millions of copies of a single fragment.
1. Many genes only have a few possible alleles. For example, humans only have a few eye colors and only four blood types. How can DNA tests definitively identify individuals when many men have brown eyes or type A blood? The way this is done is by VNTR ( variable number tandem repeats) the length varies from person to person and the base pairing also differs as well.
Experiment-Specific Questions
Part 3: Analyzing the Results
1. List the distances traveled (in mm) for the bands in the DNA Ladder in the table below.
Remember, smaller fragments travel farther than longer ones, so the top-most band will be the 1,000 bp fragments while the bottom-most band will be the 50 bp fragments.
DNA Ladder
Band
Distance (mm)
50 bp 100 bp
150 bp 200 bp
250 bp 300 bp
400 bp 500 bp
600 bp 700 bp
800 bp 900 bp
1,000 bp
2.
1. The DNA fragments
There are three specific steps required to isolate DNA from its cellular contents. The steps used to remove and expose DNA from its cell are: breaking down the food type you are using by crushing it, for example a banana or strawberries, exposing the substance to a sodium chloride (NaCl) solution, subjecting the product to detergent solution (dH2O), filtering the solution and lastly, the addition of ethanol. When beginning with a solid substance, such as a banana, crushing the substance allows for
(PCR), which isolates small fragments of DNA that have a high degree of variability from
Table 2: Consists of color extract taken from a red cabbage for a natural indicator. The pH reading that was measured by using the pH meter and the result of the pH reading to determine whether the solution was acidic or basic.
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Figure 2: As the unknown was identified as galactose or lactose and the main difference between them was that galactose was monosaccharide, while lactose was disaccharide, Barfoed’s test was performed next. As expected, all monosacharides (fructose, galactose, glucose, xylose) formed a small amount
When analyzing DNA it is important to understand it and all the chemicals that it is made of. The first thing that’s important to know is simply what DNA stands for, which is deoxyribonucleic acid. The chemical units are called nucleotides, and each nucleotide has a compound of phosphate sugar which is the backbone, and a sugar deoxyribose. The Phosphates and sugars are the same in all nucleotides but the one thing that is different would be the bases. DNA bases are cytosine, thymine, adenine, and guanine. Each base has specific partner, for example Cytosine will always pair with guanine. And Thymine will always pair with adenine.
An electrophoresis chamber is used as a way to separate DNA and their particles, based upon their sizes and charges. The shorter particles will move faster than the longer ones because the shorter particles are able to move easier through the pores of the agarose gel. The structure of DNA, also known as deoxyribonucleic acid, is a double helix. The structure is composed of nucleotides that contain important genetic information used to help organisms survive, reproduce, and develop. The information is found inside every cell of organisms. The nucleotide on the DNA is made up of a phosphate group, a sugar group, and a nitrogenous base. There are four types of nitrogenous bases: adenine (A), thymine (T), guanine (G) and cytosine (C). DNA’s information is determined by the order of these nitrogenous bases. Micropipettes can be used in the scientific field for transferring, measuring, or injecting small amounts of liquids. Micropipettes vary in sizes; they are capable of pipetting different amounts of volumes: (P20) = 0.5- 20 microL, (P200) = 20- 200 microL, and (P1000) = 100- 1000 microL. A centrifuge is a machine that is used to separate fluids with different solidities, which is constantly
Many people are not aware of what professionals are responsible for in the fields of Management Information Systems and Health Care Information Systems or what well-paying jobs are available in those fields. Some people know a small of information about them but do nor do they understand why someone would want to major in these fields. This information obtained research of these fields will help the reader become more familiar help you become more familiar with what they are, what they do, the career choices they can provide, and how these fields is very dominate in today’s job market.
Modern laboratory techniques benefits them to remove DNA from tissue samples, thereby tiny amounts of DNA from thousands of individual cells will spill together. Once the DNA is collected and purified, thereby is white, sticky substance that is somewhat translucent. In the past, Rosalind Franklin, used x-ray diffraction, which is needed by researchers to actually visualize the double helix structure of DNA. However, with a standard light microscope it's possible to see chromosome if they are in their most condensed form. Soon after, karyotype was formed through all the visualized chromosome within the cell and captured images of them to be arranged into a composite
Gel electrophoresis is a simple technique that allows us to determine the charges and molecular weights of all sorts of macromolecules. The basic theory is a simple one: more negatively charged molecules will migrate in an electric field, toward the positively charged cathode. A matrix (such as agarose or polyacrylamide) must be used to conduct heat evenly and provide an extra sieving effect. In particular, agarose gel electrophoresis is generally used to separate DNA (single-stranded, double-stranded, and supercoiled) and RNA. Since DNA is negatively charged, it migrates in an electric field toward the positively charged cathode.
A DNA marker (size standard or a DNA ladder) is loaded into the first well of the gel. The fragments in the marker are of a known length so it can be used to help approximate the size of the fragments in the samples. The prepared DNA samples are then pipetted into the remaining wells of the gel. When this is done the lid is placed on the electrophoresis tank making sure that the orientation of the gel and positive and negative electrodes is correct. To separate the fragments, the electrical current is then turned on so that the negatively charged DNA moves through the gel towards the positive side of the gel. The distance the DNA has migrated in the gel can be judged visually by monitoring the migration of the loading buffer dye. The electrical current is left on long enough to ensure that the DNA fragments move far enough across the gel to separate them, but not so long that they run off the end of the gel.
To be able to analyze our samples based on a standard curve, control samples (provided in the kit) were diluted according to the protocol from Epigentek. Each plate consisted of duplicates of each sample and controls. For each sample, 50-200 ng of DNA was used for as recommended by Epigentek. The standard absolute quantification protocol from Epigentek was modified according to recommendations from Epigentek (personal communication). Briefly, those modifications were using a multichannel pipette, using the same tips to add solutions to the wells and then a new set to remove solutions, not touching the sides of the wells with the pipette, tilting the plate to pipette solutions out, keeping a consistent timing during color changing steps and covering the plate with foil during the color changing steps. In order to decrease the percent coefficient of variation between duplicate samples, additional modifications were implemented. Specifically, for 17 of the samples, a method of splashing the solution in the plate wells into a bucket and then banging the plate on paper towels until the wells appeared dry was used instead of the pipetting the solution of the wells by tilting the plate. Absorbance readings from each plate were calculated at 450 nm using a BioTek Synergy Plate Reader and the Gen 5 Program Data Analysis Software
The relative band size of the DNA bands influences the distance that which it travels along the gel. The less base pairs within each band, the further it travels on the gel. This is confirmed in table 1 which was created by analyzing Lane 1, the molecular weight markers on agarose gel #2. The distance migrated was compared to the size of each band. Table 1 confirms this assertion as the band with 1517 bp only traveled 25 mm on the gel where as the band
After collection and sequencing, the DNA collected from human cheek cells was able to be analyzed in a variety of ways. In order to determine whether or not a significant amount of DNA was present, the first analysis done on the sample was gel electrophoresis. Figure 1 shows an image of the electrophoresis gel taken while it was being exposed to UV light. Even though both the DNA samples and the ladder were stained with ethidium bromide, a stain that fluoresces under UV light, only the ladder appeared on the gel when exposed to ultraviolet radiation. Lanes 2 and 3, the sample DNA lanes, appeared empty. This indicates that the DNA existed in small enough quantities that it could not be viewed within the agarose gel. Since the electrophoresis
To sequence, these slides are mixed with DNase and fluorescence-labelled single nucleotide. The terminator on these marked nucleotides ensures the bases are added in a one-by-one order. Slides images are taken in each cycle. In each reading site, a fluorescent signal is captured and the type of the base added are recognised. Then, the slide proceeds to the next cycle. The next base is added after the cleavage of the previous terminator. Signal from former cycle is removed to prevent interference. This process is repeated: one nucleotide is added at a time with one image taken. A sequence can then be constructed by