Dna Digestion and Electrophoresis - 728 Words

DNA DIGESTION AND ELECTROPHORESIS In this experiment we will be doing a process called as DNA digestion or also known as restriction digest. A restriction digest is a procedure used in molecular biology to prepare DNA for analysis or other processing. It is sometimes termed DNA fragmentation, scientists Hartl and Jones describe it this way: This enzymatic technique can be used for cleaving DNA molecules at specific sites, ensuring that all DNA fragments that contain a particular sequence have the same size; furthermore, each fragment that contains the desired sequence has the sequence located at exactly the same position within the fragment. The cleavage method makes use of an important class of DNA-cleaving enzymes isolated primarily†¦show more content†¦One of the most important reaction conditions which varies between different restriction enzymes is the salt concentration. Enzyme buffers are specifically formulated to provide the salt concentration for optimal enzyme activity. It is important, therefore, that the correct buffer solution is used for a particular restriction enzyme. [3] For this experiment we also made use of agarose gel electrophoresis, which takes a lot of time. Electrophoresis may be the main technique for molecular separation in todays cell biology laboratory. In spite of the many physical arrangments for the apparatus, and regardless of the medium through which molecules are allowed to migrate, all electrophoretic separations depend upon the charge distribution of the molecules being separated. Electrophoresis can be one dimensional or two dimensional. One dimensional electrophoresis is used for most routine protein and nucleic acid separations. Two dimensional separation of proteins is used for finger printing , and when properly constructed can be extremely accurate in resolving all of the proteins present within a cell. The support medium for electrophoresis can be formed into a gel within a tube or it can be layered into flat sheets. The tubes are used for easy one dimensional separations, while the sheets have a larger surface area and are better for two- dimensional separations. In electrophoresis, proteins are separated on the basis ofShow MoreRelatedIsolation, Restriction Digestion, And Electrophoresis Of Plasmid Dna1827 Words   |  8 PagesIsolation, restriction digestion, and gel electrophoresis of plasmid DNA Prathyusha Gudapati, BIOL 304, spring 2015. Abstract The purpose of the experiment was to isolate plasmid DNA, followed by restriction digestion using restriction endonucleases and then visualizing the digested fragments after subjecting to gel electrophoresis. Plasmid DNA (pSP72 DNA) was isolated from Escherichia coli KAM32 (E.coli) cultures using the QIA prep miniprep kit and then subjected to restriction digestion by EcoRI and HindIIIRead MoreMethods Of Restriction Mapping Site Essay730 Words   |  3 Pages Discussion In this experiment two methods of restriction mapping site were used i.e. double digestion method and the comparison of fragment pattern to a known sequence. In the double digestion method, the fragments produced by the enzyme Hind III andXhoI independently and concurrently were compared so that it can produce an estimation of relative postion of cleavage site The drawback of restriction mapping is that the technique cannot be directly used for eukaroyotes because of difficulties associatedRead MoreRestriction Enzyme, Alkaline Phosphatase Digestion And Gel Electrophoresis1195 Words   |  5 PagesMMR Report 1.3 Restriction Enzyme, Alkaline phosphatase Digestion †¨and Gel Electrophoresis By Naga Srilekha Somu Chemistry - 429 Spring 2016 Western Illinois University Materials and Equipment: Pure plasmid pET28a, amplified 2-alcohol dehydrogenase gene (a PCR product), 10x bovine serum albumin, 10x neutralization buffer, EcoRI, nuclease free water, pET28a plasmid digested with EcoRI, calf intestinal alkaline phosphatase, agarose gel (1% agarose + 0.3ÃŽ ¼L ethidium bromide), 1x TAE bufferRead MoreA Research Study On Scar Markers1635 Words   |  7 PagesSCAR markers are PCR based primers that represent genomic DNA fragments at genetically defined loci, that are identified by PCR amplification using sequence specific oligonuceotide primers (Paran and Michelmore, 1993; Me Dermott et al., 1994). Inception of SCARs involves cloning the amplified products of arbitrary marker techniques and then sequencing the 2 ends of the cloned products. The sequence s therefore used to design specific primer pairs of 15-30 bp which will amplify single major bandsRead MoreEssay on Using PCR and Gel Electrophoresis to Determine Genotype583 Words   |  3 PagesUsing PCR and Gel Electrophoresi s to Determine Genotype In certain situations, it is necessary to identify DNA retreived from a sample. When there is a small sample in need of identification, Polymerase Chain Reactions are used to multiply the DNA in the sample in to many identical samples. The DNA retrieved from the reaction can then be imported into an aparatus using gel electrophoresis to compare the sample of DNA to other samples. In our experiment we learned the how to replicate tinyRead MoreOptimization of Asymmetric PCR for Generation of a Single Stranded DNA Library690 Words   |  3 PagesAptamers are short DNA or RNA oligonucleotides with high, specific affinity to a special target. The name was originated from aptus that means to fit and meros that shows the polymer identity of oligonucleotides (1, 2). Aptamer characteristics provide prominent potential applications in multiple fields.These nucleic acid ligands are completely generated through in vitro process for a wide range of targets from small molecules and ions to large proteins and cells and even whole organism or tissueRead More1.3.1 response Essay624 Words   |  3 Pagesï » ¿ Activity 1.3.1: Student Response Sheet PART A- Restriction Enzymes Restriction enzymes are a tool that allows us to pinpoint human identity down to single differences in our DNA. Work through the following simulation so you can see these molecular scissors in action. Find out more about restriction enzymes by viewing the animation and reading the article listed below. DolanDNALearningCenter: Restriction Enzymes http://www.dnalc.org/ddnalc/resources/restriction.html Access ExcellenceRead MorePlant Viruses : A Large, Unique Family Of Plants Viruses1458 Words   |  6 Pagesworld. [1, 2] Geminiviruses consist of four genera, Mastrevirus, Curtovirus, Begomovirus, and Topocuvirus based on host ranges, vector specificities, and genome organizations. Characteristics of geminiviruses include their circular, single-stranded DNA genome and geminate-shaped virus particles. These viruses-- exhibit both prokaryotic and eukaryotic features-- replicate in the nuclei of the host cell and depend on host machineries for transcription [1]. The majority of the Old World begomovirusesRead MorePcr Rflp Report : Pcr1166 Words   |  5 PagesPCR-RFLP Report PCR What is it: The Polymerase Chain Reaction is a method that uses the capability of DNA polymerase to synthesize to new DNA strands which are matching to the template strand. A primer needs to be added to the first nucleotide due to the fact that DNA polymerase only can add a nucleotide only onto a 3 -OH group that already exists. Because of this condition, we are able to define a chosen region of template sequence which we can then generate millions to billions of copies. ThisRead MoreSite Directed Mutagenesis ( Sdm ) Technique942 Words   |  4 PagesSite-directed mutagenesis (SDM) technique is commonly used to induce desired change in DNA plasmid sequence by mutation, insertion or deletion with oligonucleotide primers (1). This SDM usually cooperate with ploymerase chain reaction (PCR) as to amplify the concentration of mutated template (2). PCR, a temperature-based cycle reaction, is completed with three initial steps including denaturing the DNA template, anneal the mutated oligonucleotide primers and elongating the mutated primer with ploymerase