Gel Electrophoresis Lab

Introduction: All organisms have DNA (Deoxyribonucleic Acid) in the nuclei of their cells. The aim of this experiment is to extract this DNA using common household detergent to breakdown the cellular membrane then through the use of ethanol to extract the DNA from the Kiwi and Strawberry fruit. After the DNA has been extracted the DNA will be visualised by agarose gel electrophoresis which is done by adding a dye to the extracted DNA then being placed in the gel, once the negatively charged DNA has been placed in the gel it will slowly move towards the positively charged end (anode) then is viewed under an ultraviolet light then compared to the control of known lengths of DNA to show the size of the unknown DNA samples.

Isolation of DNA is mainly done by adding different other digestive enzyme which reacts with constituents in the sample encountered at the crime scene, other than DNA. The different enzymes are proteases which breakdowns amino acids and proteins RNAase which digest RNA particles thus leaving only DNA in the test-tube. Ice cold ethanol is added to this homogenate to precipitate DNA. The DNA can be isolated slowly stirring a glass rod which is known as spooling.

In this lab, evidence DNA and the DNA samples of two different suspects were available. Two restriction enzyme digests were conducted to cut the samples into smaller pieces. An increase in the number of restriction enzyme digests increases accuracy and decreases the likelihood of false positives. This is crucial in forensic DNA analysis. Finally, the DNA was separated using Electrophoresis. Agarose Gel Electrophoresis is the most effective way of separating DNA fragments of varying sizes and charge (Lee, 2012). Agarose gel is ideal since during gelation, the agarose molecules are held together by hydrogen bonds which form pores uniformly throughout the gel giving it its molecular sieving properties (Lee, 2012).The DNA to be separated is inserted into the pre-cast depressions/wells in the gel (Lee, 2012). The gel is then inserted into a chamber and an electrically conductive buffer solution is added until the gel is completely submerged (Lee, 2012). Positive and negative electrodes are connected to each end of the chamber and an electric current is applied (Lee, 2012). Since the phosphate backbone of the DNA molecule is negatively charged, the DNA will be repelled from the negative electrode and will move toward the positive electrode (Lee, 2012). DNA bands will be visible in the gel with varying distances (Lee, 2012). Smaller fragments will move farther throughout the porous gel.

Implementation of Gel Testing for Antibody Screening and Identification in a Community Hospital, a 3-Year Experience

Gel electrophoresis proving to give a very positive and forward moving impact in the science field, has many advantages. For one this scientific technique caters a clear link

Gel electrophoresis is used to separate and to analyse macromolecules such as DNA, RNA and proteins and their fragment based on their length, to estimate the size of DNA. The agarose gels submerged into electrophoresis buffer. The macromolecule (need to be added with ethidium bromide(dye) so that the DNA will appear to be fluorescent in red-orange colour under ultraviolet light) are separated by applying an electric field to move the charged molecules through an agarose matrix and the molecules are separated by size in the agarose gel matrix. The DNA length are measured in bp. As the DNA molecules move in the gel, it takes up the dye. A number of factors can affect the migration of nucleic acids: the dimension of the gel pores (gel concentration),

DNA fingerprinting is used as a forensic skill in criminal investigations to identify an unidentified person or whose identity needs to be confirmed. A tissue sample is obtained and the DNA is extracted. Fragments of different lengths are made by cutting up the DNA. The fragments are split up and visualized in some way and is compared with another sample. In our lab, we practiced a method called Gel electrophoresis a technique commonly used in labs and clinics to separate charged molecules like RNA and DNA according to their size. DNA is negatively charged providing an electric current to the gel, DNA will travel towards the positively charged electrodes. The Short strands of Deoxyribose move quickly through the gel template than longer strands

Agarose gel electrophoresis AGE) is a method used to separate DNA molecules into fragments. This can be done by moving negatively charged nucleic acid molecules through an agarose matrix with an electrostatic field (electrophoresis). The shorter molecules able to move faster and migrate farther than longer

The article describes using the method of gel electrophoresis to separate DNA fragments to identify and analyze certain sequences. The DNA fragments were broken down using a restriction enzyme from Haemophilus influlenzae, allowing the researchers to see that these restriction enzymes can be used to study DNA and its structure. The first step of the gel electrophoresis was to set the gel matrix. Agarose was used to separate DNA molecules, and acrylamide was used to separate proteins. The gel starts off as a liquid, which was poured into a molding tray. A comb was placed in the liquid matrix so that when the matrix solidified, wells formed to load samples in them. After the gel solidified, the mold was removed and placed

(http://www.banglajol.info/index.php/AKMMCJ/article/viewFile/13682/9836). This method was used as it amplifies the DNA, which is required. Secondly, Agrose Gel Electrophoresis was used. “This laboratory technique is used to separate fragments of DNA by a charge.” (http://www.biotecharticles.com/Applications-Article/Agarose-Gel-Electrophoresis-805.html)

Gel purification allows us to isolate and purify DNA fragments based on size. The DNA bands from the agarose gel are cut out and purified using some procedure. The band was extracted by using the QIAgen gel extraction protocol. The DNA bands after purification for CD01 and CD02 were shown in Figure 4.3. This steps also considered to be vital components in molecular biology technique as it can help determine the success or failure of the downstream

Agarose gel electrophoresis is a common technique that is used to separate and identify unknown DNA molecules according to their sizes and structures. Agarose gel enables this with the pores it contains. These pores act as a sieve, and allows the DNA molecules to separate by size and conformation; the larger and more linear the molecules are, the slower they will travel down the gel. These DNA molecules are able to travel due to their negative property caused by their phosphate groups. Once the gel is running in electric fields, the negatively charged DNA molecules travel to the positive end of the electrode. Once running the gel is finished, the gel is compared with a known standard to determine the size and concentration of the DNA in the

This process utilizes yellow or green gelatin or cellophane filter recorded on a camera at an illumination of 300-nm UV light. Applications of Electrophoresis of Nucleic Acid: • Agarose Gel Electrophoresis is employed in restriction mapping of cloned DNA i.e. to estimate the size of DNA fragments digested with restriction enzymes. • It has application in molecular genetics & genetic fingerprinting of PCR products. • It is used to separate genomic DNS (Southern Blot) and RNA (Northern Blot).

Separation of strands occurs via as a electrical gradient is applied to a gel matrix in which DNA samples are loaded. Due to the slight negative charge of the DNA strands, they will move towards the positive charge. Larger molecules move more slowly by this method, so we can find a DNA fingerprint4 to compare with other samples of DNA .

Since the charge per unit length (owing to the phosphate groups) in any given fragment of DNA is the same, all DNA samples should move towards the anode with the same mobility under an applied electrical field. However, separation in agarose gels is achieved because of resistance to their movement caused by the gel matrix. The largest molecules will have the most difficulty passing through the gel pores (very large molecules may even be blocked