Free

Agarose Gel Electrophoresis of DNA - Term Paper Example

Comments (0) Cite this document
Summary
Agarose Gel Electrophoresis of DNA Introduction Electrophoresis refers to a method used to separate and purify macromolecules, mostly nucleic acids and proteins, which differ in conformation, size, or charge…
Download full paperFile format: .doc, available for editing
GRAB THE BEST PAPER96.2% of users find it useful
Agarose Gel Electrophoresis of DNA
Read TextPreview

Extract of sample "Agarose Gel Electrophoresis of DNA"

Agarose Gel Electrophoresis of DNA Introduction Electrophoresis refers to a method used toseparate and purify macromolecules, mostly nucleic acids and proteins, which differ in conformation, size, or charge. It is among the widely used techniques in molecular biology and biochemistry. Gel electrophoresis refers to the method of analysis and separation of macromolecules (proteins, RNA, and DNA) and their respective fragments, depending on their charge and size. Gel electrophoresis is applied in clinical chemistry to separate proteins by size and/or charge and in molecular biology and biochemistry to separate RNA and DNA fragments by length, separating proteins by charge, or estimating the size of RNA and DNA fragments. To separate nucleic acid molecules, an electric field is applied to move molecules with negative charges through an agarose matrix. The short molecules move fast hence migrating farther compared to the longer ones. This is due to the molecule’s ability to easily migrate via the pores of the gel. A charge separates proteins in agarose due to the pores of the gel being too large to sieve them (proteins). Gel electrophoresis is also used in the separation of nanoparticles (Colorado State University, 1). Agarose Gel Electrophoresis of DNA in Context Agarose is a polysaccharide which is extracted from seaweed. Typically it is used at 0.5% to 2% concentrations. A high agarose concentration makes the gel stiffer, and vice versa. Agarose gels are easy to prepare and non toxic. Agarose powder is simply mixed with a buffer solution then melted by heating, and then the gel is poured. Agarose gels have a low resolving power, but relatively a large separation range. DNA fragments from around 200 to 50,000 base pair can be separated by varying agarose concentration using electrophoretic techniques. Agarose gels are easily handled and cast than other matrices since the gel setting is not a chemical but a physical change with an easy recovery of samples (Colorado State University, 1). Agarose gel electrophoresis is used in the separation of DNA fragments which range from 50 base pair up to several megabases by use of specialized apparatus. The agarose percentage in the gel determined the distance between DNA bands of a certain length. Majority of the agarose gels comprise of between 0.7% (a good resolution for large DNA fragments of 5–10kb) to 2% (a good resolution for 0.2–1kb DNA fragments) of agarose which is dissolved in a buffer of electrophoresis. Low percentage gels are extremely weak and can break when lifted. High percentage gels are mostly brittle and unevenly set. Agarose gels lack uniformity in pore size. However, they are optimum for electrophoresis of proteins which are larger than 200 kDa (National Center for Biotechnology Information, 1). Agarose gel electrophoresis is so far the most effective method of separating varying sizes of DNA fragments with a range of 100 bp and 25 kb. Agarose is separated from the seaweed Gracilaria and genera Gelidium and contains repeated agarobiose (D- and L-galactose) units. Agarose polymers non-covalently associate during gelation to form a bundles network. The size of the pores determines the molecular sieving properties of a gel. To separate DNA, it is placed into pre-cast wells inside the gel and then a current is applied. The DNA molecules phosphate backbone is negatively charged, so when it is placed within an electric field, the DNA fragments migrate to the anode which is positively charged. Since DNAs mass/charge ratio is uniform, its molecules within an agarose gel are separated by size in a certain pattern such that the traveled distance is inversely proportional to its molecular weight log. The migration rate of a DNA molecule within a gel is determined by: DNA molecule size; concentration of agarose; DNA conformation; applied voltage; ethidium bromide presence; electrophoresis buffer; and agarose type. After the separation, DNA molecules are stained with an appropriate dye and therefore visible under ultra-violet light (U.S National Library of Medicine, 1). Conclusion The adoption of agarose gel electrophoresis has revolutionized DNA separation. Before to the adoption, DNA used to be separated by the use of sucrose density gradient centrifugation. This only gave approximations by size. By adopting agarose gel electrophoresis, scientists are able to understand the mechanisms behind the separation of DNA fragments within a gel matrix. The DNA molecule conformation helps them to determine the mobility via a gel matrix. With the identification of an appropriate concentration for the agarose solution; preparation of an the agarose gel for electrophoresis; setting up the power supply and apparatus; selecting the appropriate voltage to separate DNA fragments; and using ethidium bromide to allow for DNA bands visualization, scientists are able to determine the different sizes of the separated DNA fragments. Works Cited Colorado State University. “Agarose Gel Electrophoresis of DNA.” Colostate.edu. 15 Jan. 2000. Web. 05 April. 2013. Colorado State University. “Principles of Gel Electrophoresis.” Colostate.edu. 15 Jan. 2000. Web. 05 April. 2013. National Center for Biotechnology Information. “Agarose gel electrophoresis for the separation of DNA fragments.” Ncbi.nlm.nih.gov 20 April. 2012. Web. 05 April. 2013. U.S National Library of Medicine. “Agarose gel electrophoresis for the separation of DNA fragments.” Nlm.nih.gov. 20 April. 2012. Web. 05 April. 2013. Read More
Cite this document
  • APA
  • MLA
  • CHICAGO
(“Agarose Gel Electrophoresis of DNA Term Paper Example | Topics and Well Written Essays - 500 words”, n.d.)
Retrieved from https://studentshare.org/biology/1472674-agarose-gel-electrophoresis-of-dna
(Agarose Gel Electrophoresis of DNA Term Paper Example | Topics and Well Written Essays - 500 Words)
https://studentshare.org/biology/1472674-agarose-gel-electrophoresis-of-dna.
“Agarose Gel Electrophoresis of DNA Term Paper Example | Topics and Well Written Essays - 500 Words”, n.d. https://studentshare.org/biology/1472674-agarose-gel-electrophoresis-of-dna.
  • Cited: 0 times
Comments (0)
Click to create a comment or rate a document

CHECK THESE SAMPLES OF Agarose Gel Electrophoresis of DNA

DNA

...chromatids separate, producing haploid cells with unreplicated chromosomes. Ideally, 4 daughter cells are produced per meiosis of a –gonium, and this is what happens in the production of sperm cells. However, in the case of female gamete formation, 2 daughter cells (1 from meiosis I and 1 from meiosis II), only 1 oocyte is produced from a cycle of meiosis (Campbell and Reece, 2002). DNA replication occurs in preparation for cell division How does DNA replicate? A part of the double-stranded DNA (dsDNA) unwinds, allowing DNA polymerase and DNA ligase to get into what is known as the replication bubble. The DNA polymerase adds the...
4 Pages(1000 words)Essay

DNA

...these proteins and biomolecules was unique. To analyze this DNA, criminal lab technicians use a technique called gel electrophoresis to filter DNA by size. DNA is broken down into uniquely sized fragments based on that person’s unique DNA sequence (Gel). These DNA fragments are then pulled through a gel by electricity at differing rates based on size. Because these fragments are uniquely sized, a picture of where the fragments lie after a given time should show fragments uniquely drawn along differing points on the gel. When compared to a sample of an already identified...
3 Pages(750 words)Essay

Agarose Gel Electrophoresis of DNA

...? Agarose Gel Electrophoresis of DNA DNA is the double stranded helical structure which carries the genetic information. The study of the DNA molecule will give us the details about life. DNA molecules can be extracted from the cell using the extraction techniques and they are then quantified using the agarose gel electrophoresis. (Westermeier 2006). Agarose gel electrophoresis is the most common and easiest technique to separate the DNA fragments. Agarose is a polymer that forms helical linking strands between the molecules and they are held together by hydrogen bonds. Based on the concentration of the agarose in the gel, the pore size also varies. The DNA fragment is separated in the Agarose based on the pore size... (that is the...
5 Pages(1250 words)Lab Report

Protein Extraction and Gel Electrophoresis

... Proteomics and Gel Electrophoresis Abstract Proteomic is generally defined as the direct analysis of proteins in terms of their presence and relative abundance. Gel electrophoresis is a significant methodology employed for extraction of proteins in proteome analysis. The following is a report on an experiment conducted to determine the concentration of proteins in two samples using the Bradford assay and separating the proteins using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It involved three steps, that is, protein extraction, protein quantitation, and gel electrophoresis of two...
5 Pages(1250 words)Lab Report

DNA Restriction and Electrophoresis

...III.Each of these enzymes has at least five or more restriction sites on the chromosome and it consequently produces six or more restriction fragments of different lengths. The DNA strands are then immersed in a well of 0.8% agarose gel, which is the chosen medium.An electrical field, is applied to the gel evenly which makes the DNA fragments move faster from their original placement towards the positively charged electrode. The smaller DNA fragments travel faster through the gel than the larger ones. The gel actually acts like a sieve. Depending on the restriction enzyme used the DNA...
4 Pages(1000 words)Lab Report

Conventional PCR using agarose gel electrophoresis detection

...Conventional PCR using agarose gel electrophoresis detection According to Lewis “Agarose gel electrophoresis is the easiest and commonest way of separating and analyzing DNA”. Conventional techniques used to detect mycoplasma involve culturing samples on selective media, which needs at least a week to obtain the results (HD BIOSCIENCES Co., LTD 2). The reason to add gel in this method is to either quantify the DNA or separate the particular band of DNA. With the addition of edhidium bromide, DNA becomes visible. This binds strongly to DNA by intercalating between the bases and is fluorescent meaning that it absorbs invisible UV light and transmits the energy as visible orange light (Lewis). While addition of gel, the care... the bromophenol...
2 Pages(500 words)Essay

DNA

...DNA DNA represents the genetic material within a cell that provides the basic blueprint for living cells, somewhat like a program for creating biological structures. Though DNA has received much attention in the media in recent years, it is important to note that the discovery and implementation of most DNA techniques only occurred as recently as the middle of the twentieth century. Understanding of the history, structure, and function of DNA is critical to modern science. Though DNA had long been known to exist is the cell, the structure and importance of the compound was not fully understood until 1953 when Francis Crick and James...
3 Pages(750 words)Term Paper

How to make and run an agarose gel (DNA Electrophoresis)

...Agarose Gel Electrophoresis of DNA For isolation of genomic DNA, agarose gel electrophoresis is done which is based on the principle that on disruption and lysis of cell components, followed by removal of proteins and sugar, nucleic acids can be recovered which can be observed through a UV transilluminator on the agarose gel due to prior staining by ethidium bromide. Materials: Agarose powder, Electrophoresis buffer (TAE), gel casting tray, Ethidium bromide, Comb, Sample, Loading buffer, UV transilluminator. Method:...
1 Pages(250 words)Article

ELECTROPHORESIS

.... A tiny well is cut through the gel through which analytes are introduced. Agarose, which is a less polar portion of agar, eliminates the effects of electroendosmosis and acts as a molecular sieve for large molecules such as proteins and DNA. Advantages of ElectrophoresisElectrophoresis is a relatively inexpensive technique as simple equipment is used. The gels are available commercially and several samples can be run concurrently on the same gel. The technique is also versatile and can be combined with isoelectric focusing to form two-dimensional electrophoresis, which permits the resolution of...
6 Pages(1500 words)Essay

Size Exclusion Chromatography- Protein Separation Polyacrylamide Gel Electrophoresis

...molecular weights incorporated in this analysis. These standards were used to plot the calibration curve. Table 1: List of standards and their Molecular weights Elution fraction of both the standards and unknown samples were run through SDS-PAGE gel electrophoresis and their retention times calculated as shown in the table below. Table 2: Retention of known standards and unknown. Retention time of standards Retention Time of unknowns Band no. Retention time unknown no. Retention time 1 0.190476 2 0.247619 2 0.209524 3 0.590476 3 0.266667 4 0.607619 4 0.361905 5 0.52 5 0.419048 6 0.495238 6 0.533333 7 0.209524 The retention times, which is equivalent to elution volume (Ve) of each protein was plotted as a...
1 Pages(250 words)Lab Report
sponsored ads
We use cookies to create the best experience for you. Keep on browsing if you are OK with that, or find out how to manage cookies.

Let us find you another Term Paper on topic Agarose Gel Electrophoresis of DNA for FREE!

Contact Us