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      "body": "<html>\n<p>&nbsp;&nbsp;<strong>Introduction</strong><br>\nThe search for a more efficient means of extracting DNA of both higher quality and yield<br>\nhas lead to the development of a variety of protocols, however the fundamentals of DNA<br>\nextraction remains the same. DNA must be purified from cellular material in a manner<br>\nthat prevents degradation. Because of this, even crude extraction procedures can still be<br>\nadopted to prepare a sufficient amount of DNA to allow for multiple end uses.<br>\nDNA extraction from plant tissue can vary depending on the material used. Essentially<br>\nany mechanical means of breaking down the cell wall and membranes to allow access to<br>\nnuclear material, without its degradation is required. For this, usually an initial grinding<br>\nstage with liquid nitrogen is employed to break down cell wall material and allow access<br>\nto DNA while harmful cellular enzymes and chemicals remain inactivated. Once the<br>\ntissue has been sufficiently ground, it can then be resuspended in a suitable buffer, such<br>\nas CTAB. In order to purify DNA, insoluble particulates are removed through<br>\ncentrifugation while soluble proteins and other material are separated through mixing<br>\nwith chloroform and centrifugation. DNA must then be precipitated from the aqueous<br>\nphase and washed thoroughly to remove contaminating salts. The purified DNA is then<br>\nresuspended and stored in TE buffer or sterile distilled water. This method has been<br>\nshown to give intact genomic DNA from plant tissue. To check the quality of the<br>\nextracted DNA, a sample is run on an agarose gel, stained with ethidium bromide, and<br>\nvisualised under UV light.</p>\n<p><img src=\"https://www.frontiersin.org/files/Articles/91373/fmicb-05-00286-r2/image_m/fmicb-05-00286-g001.jpg\" width=\"892\" height=\"717\"/><br>\n<strong>Materials</strong><br>\nCTAB buffer<br>\nMicrofuge tubes<br>\nMortar and Pestle<br>\nLiquid Nitrogen<br>\nMicrofuge<br>\nAbsolute Ethanol (ice cold)<br>\n70 % Ethanol (ice cold)<br>\n7.5 M Ammonium Acetate<br>\n55o C water bath<br>\nChloroform : Iso Amyl Alcohol (24:1)<br>\nWater (sterile)<br>\nAgarose<br>\n6x Loading Buffer<br>\n1x TBE solution<br>\nAgarose gel electrophoresis system<br>\nEthidium Bromide solution<br>\n<strong>CTAB buffer 100ml</strong><br>\n2.0 g CTAB (Hexadecyl trimethyl-ammonium bromide)<br>\n10.0 ml 1 M Tris pH 8.0<br>\n4.0 ml 0.5 M EDTA pH 8.0 (EthylenediaminetetraAcetic acid Di-sodium salt)<br>\n28.0 ml 5 M NaCl<br>\n40.0 ml H2O<br>\n1 g PVP 40 (polyvinyl pyrrolidone (vinylpyrrolidine homopolymer) Mw 40,000)<br>\nAdjust all to pH 5.0 with HCL and make up to 100 ml with H2O.<br>\n<strong>1 M Tris pH 8.0</strong><br>\nDissolve 121.1 g of Tris base in 800 ml of H2O. Adjust pH to 8.0 by adding 42 ml of<br>\nconcentrated HCL. Allow the solution to cool to room temperature before making the<br>\nfinal adjustments to the pH. Adjust the volume to 1 L with H2O. Sterilize using an<br>\nautoclave.<br>\n<strong>5x TBE buffer</strong><br>\n54 g Tris base<br>\n27.5 g boric acid<br>\n20 ml of 0.5M EDTA (pH 8.0)<br>\nMake up to 1L with water.<br>\nTo make a 0.5x working solution, do a 1:10 dilution of the concentrated stock.<br>\n<strong>1% Agarose gel</strong><br>\n1 g Agarose dissolved in 100 ml TBE<br>\n<strong>Procedure</strong><br>\n- Grind 200 mg of plant tissue to a fine paste in approximately 500 μl of CTAB buffer.<br>\n- Transfer CTAB/plant extract mixture to a microfuge tube.<br>\n- Incubate the CTAB/plant extract mixture for about 15 min at 55o C in a recirculating<br>\nwater bath.<br>\n- After incubation, spin the CTAB/plant extract mixture at 12000 g for 5 min to spin<br>\ndown cell debris. Transfer the supernatant to clean microfuge tubes.<br>\n- To each tube add 250 μl of Chloroform : Iso Amyl Alcohol (24:1) and mix the<br>\nsolution by inversion. After mixing, spin the tubes at 13000 rpm for 1 min.<br>\n- Transfer the upper aqueous phase only (contains the DNA) to a clean microfuge<br>\ntube.<br>\n- To each tube add 50 μl of 7.5 M Ammonium Acetate followed by 500 μl of ice cold<br>\nabsolute ethanol.<br>\n- Invert the tubes slowly several times to precipitate the DNA. Generally the DNA can<br>\nbe seen to precipitate out of solution. Alternatively the tubes can be placed for 1 hr at<br>\n-20 o C after the addition of ethanol to precipitate the DNA.<br>\n- Following precipitation, the DNA can be pipetted off by slowly rotating/spinning a<br>\ntip in the cold solution. The precipitated DNA sticks to the pipette and is visible as a<br>\nclear thick precipitate. To wash the DNA, transfer the precipitate into a microfuge<br>\ntube containing 500 μl of ice cold 70 % ethanol and slowly invert the tube. Repeat.<br>\n((alternatively the precipitate can be isolated by spinning the tube at 13000 rpm for a<br>\nminute to form a pellet. Remove the supernatant and wash the DNA pellet by adding<br>\ntwo changes of ice cold 70 % ethanol )).<br>\n- After the wash, spin the DNA into a pellet by centrifuging at 13000 rpm for 1 min.<br>\nRemove all the supernatant and allow the DNA pellet to dry (approximately 15 min).<br>\nDo not allow the DNA to over dry or it will be hard to re-dissolve.<br>\n- Resuspend the DNA in sterile DNase free water (approximately 50-400 μl H2O; the<br>\namount of water needed to dissolve the DNA can vary, depending on how much is<br>\nisolated). RNaseA (10 μg/ml) can be added to the water prior to dissolving the DNA<br>\nto remove any RNA in the preparation (10 μl RNaseA in 10ml H2O).<br>\n- After resuspension, the DNA is incubated at 65o C for 20 min to destroy any DNases<br>\nthat may be present and store at 4o C.<br>\n- Agarose gel electrophoresis of the DNA will show the integrity of the DNA, while<br>\nspectrophotometry will give an indication of the concentration and cleanliness.<br>\n<strong>DNA quality confirmation</strong><br>\n– Prepare a 1 % solution of agarose by melting 1 g of agarose in 100 mL of 0.5x TBE<br>\nbuffer in a microwave for approximately 2 min. Allow to cool for a couple of<br>\nminutes then add 2.5 μl of ethidium bromide, stir to mix.<br>\n– Cast a gel using a supplied tray and comb. Allow the gel to set for a minimum of 20<br>\nmin at room temperature on a flat surface.<br>\n– Load the following into separate wells<br>\no 10 μL 1kb ladder<br>\no 5 μL sample + 5 μL water + 2 μL 6x Loading Buffer<br>\n– Run the gel for 30 min at 100 V<br>\n– Expose the gel to UV light and photograph (demonstration)<br>\n– Confirm DNA quality, presence of a highly resolved high molecular weight band<br>\nindicates good quality DNA, presence of a smeared band indicates DNA<br>\ndegredation.<br>\n</p>\n</html>",
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      "author": "genetics.life",
      "body": "i can already read it bro thx",
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      "author": "cheetah",
      "body": "Hi! I am a robot. I just upvoted you! I found similar content that readers might be interested in:\nhttp://www.nbpgr.ernet.in/Portals/6/DMX/GENOMIC_RESOURCES/DNA%20extraction-Comparison%20of%20methodologies.pdf",
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      "body": "<html>\n<p>&nbsp;&nbsp;<strong>Principle</strong><br>\nGood quality DNA is a prerequisite for all experiments of DNA manipulation. All plant DNA<br>\nextraction protocols comprise of the basic steps of disruption of the cell wall, cell membrane<br>\nand nuclear membrane to release the DNA into solution followed by precipitation of DNA<br>\nwhile ensuring removal of the contaminating biomolecules such as the proteins,<br>\npolysaccharides, lipids, phenols and other secondary metabolites. This is brought about by<br>\ndisruption of the tissue in a mortar and pestle aided by liquid nitrogen and the various<br>\ncomponents of the homogenization or extraction buffer followed the precipitating and<br>\npurification methods employed. Since DNA can be extracted from various types of tissues<br>\nsuch as seedlings, leaves, cotyledons, seeds, endosperm, tissue culture callus, roots etc., the<br>\ntissue type along with the concentration of DNA finally required determine the methodology<br>\nof DNA extraction to be followed by the experimenter. The most commonly used basic plant<br>\nDNA extraction protocols are those of Dellaporta et al. 1983 and Saghai Maroof et al.,1984<br>\nalong with many others that are modifications of the components of these protocols to suit<br>\na particular tissue type or downscaling them for miniprep. In addition to these basic<br>\nprotocols, a panorama of DNA isolation kits based on either anion exchange<br>\nchromatography or silica gel membranes are available commercially.</p>\n<p><img src=\"https://image.slidesharecdn.com/281lec29moltech1-150820155016-lva1-app6892/95/281-lec29-moltech1-31-638.jpg?cb=1440085956\" width=\"638\" height=\"479\"/><br>\n<strong>Components</strong><br>\nThe role various components of DNA extraction protocol is as follows:<br>\nA. The extraction buffer<br>\nThis includes a detergent such as cetyl trimethyl ammonium bromide(CTAB) or SDS<br>\nwhich disrupts the membranes, a reducing agent such as B mercaptoethanol which<br>\nhelps in denaturing proteins by breaking the disulfide bonds between the cysteine<br>\nresidues and for removing the tanins and polyphenols present in the crude extract, a<br>\nchelating agent such as EDTA which chelates the magnesium ions required for<br>\nDNase activity , a buffer which is almost always Tris at pH 8 and a salt such as<br>\nsodium chloride which aids in precipitation by neutralizing the negative charges on<br>\nthe DNA so that the molecules can come together.<br>\nB. Phenol chloroform extraction<br>\nNucleic acid solutions commonly contain undesirable contaminants that are chiefly<br>\nmade of proteins. A classic method of purifying is phenol –chloroform extraction by<br>\nwhich the the nucleic acid solution is extracted by successively washing with a<br>\nvolume of phenol(pH 8.0); a volume of phenol: chloroform: isoamyl alcohol (25: 24:<br>\n2<br>\n1) and chloroform: isoamyl alcohol ( 24:1). Centrifugation is performed<br>\nintermittently and the upper aqueous phase is transferred to a new tube while<br>\navoiding the interphase. The contaminants are denatured and and accumulate in the<br>\norganic phase or in the marginal layer between the two phases and the nucleic acids<br>\nare preserved in the aqueous phase. Another way of removing proteins is by using<br>\nthe enzyme proteinase K which however again is denatured by phenol via phenol<br>\nchloroform extraction.<br>\nC. Precipitation of nucleic acids<br>\nAlcohol precipitation is the most commonly used method for nucleic acid<br>\nprecipitation. This requires diluting the nucleic acid with a monovalent salt , adding<br>\nalcohol to it and mixing gently. The nucleic acid precipitated spontaneously and can<br>\nbe pelleted by centrifugation. The salts and alcohol remnants are removed by<br>\nwashing with 70% alcohol. The most commonly used salts include sodium acetate<br>\npH 5.2(final volume 0.3M), sodium chloride (final concentration 0.2M), ammonium<br>\nacetate (2- 2.5M), lithium chloride (0.8M) and potassium chloride. Ethanol (twice<br>\nthe volume) or isopropanol ( two thirds volume) are the standard alcohols used for<br>\nnucleic acid precipitation.<br>\nD. Resuspending DNA<br>\nThe nucleic acid pellet can be resuspended in either sterile distilled water or TE(10<br>\nmM Tris:1mM EDTA)<br>\nE. Purification of DNA<br>\nThe DNA is purified by incubating the nucleic acid solution with RNase A (10mg/ml)<br>\nat 37°C and reprecipitation following phenol: chloroform extraction to remove the<br>\nRNase.<br>\n</p>\n</html>",
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      "body": "<html>\n<p>Carbohydrates have several roles in living organisms, including energy transportation, as well as being structural components of plants and arthropods.</p>\n<p><strong>Carbohydrates and nutrition</strong></p>\n<p>Bread, pasta, beans, potatoes, bran, rice, and cereals are carbohydrate-rich foods. Most carbohydrate-rich foods have high starch content. Carbohydrates are the most common source of energy for most organisms, including humans.</p>\n<p><img src=\"http://powerpictures.crystalgraphics.com/photo/foods_highest_in_carbohydrates_cg2p07767089c_th.jpg\" width=\"275\" height=\"196\"/></p>\n<p><br></p>\n</html>",
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      "author": "muscleroast.com",
      "body": "Strawberries are one of the healthiest foods on earth, and people don't know that.",
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      "author": "genetics.life",
      "body": "<html>\n<p>&nbsp;Strawberries grow in bushes and are delicious seasonal <a href=\"https://www.organicfacts.net/health-benefits/fruit?utm_source=internal&amp;utm_medium=link&amp;utm_campaign=smartlinks\">fruits</a> that also boost your health. &nbsp;</p>\n<p><strong>&nbsp;Health Benefits Of Strawberry:</strong></p>\n<p>&nbsp;Eye Care</p>\n<p>Boosts Immunity</p>\n<p>&nbsp;Boosts Brain Function</p>\n<p>&nbsp;Prevents Cancer</p>\n<p>Treats Arthritis and Gout</p>\n<p>&nbsp;Reduces Hypertension</p>\n<p>&nbsp;Improves Heart&nbsp;Function</p>\n<p>&nbsp;Weight Loss</p>\n<p>&nbsp;Prevents Birth Defects</p>\n<p>&nbsp;Reduces Inflammation</p>\n<p><img src=\"http://d13z1xw8270sfc.cloudfront.net/origin/450986/1499718462277_strawberry.jpg\" width=\"885\" height=\"470\"/></p>\n</html>",
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      "author": "genetics.life",
      "body": "<html>\n<p>As bright as the rising sun,</p>\n<p>You have lit up every day</p>\n<p>As dark as night without moon</p>\n<p>Are days where you away.</p>\n<p>when I'm with you life free</p>\n<p>And the world is without falt;</p>\n<p>In your hand you hold the key,</p>\n<p>The key may heart's great value</p>\n<p><img src=\"http://www.a-w-i-p.com/media/blogs/poetry/Poetry4/sun_sunrise_autumn_73.jpg\"/></p>\n<p><br></p>\n</html>",
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      "body": "<html>\n<p>QUAID-E-AZAM MUHAMMAD ALI JINNAH is the founder of Pakistan.</p>\n<p><img src=\"http://blog.chughtaimuseum.com/wp-content/uploads/Quaid-e-Azam.jpg\"/></p>\n</html>",
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      "author": "talatawan",
      "body": "Nice and informative bro....\nUpvote you please review my post",
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      "body": "<html>\n<p><strong>Plant introduction</strong> consists of taking a genotype or a group of genotypes of plants into new environments where they were not being grown before. Introduction may involve new varieties of a crop already grown in the area, wild relatives of the crop species or a totally new crop species.</p>\n<p><img src=\"https://encrypted-tbn0.gstatic.com/images?q=tbn:ANd9GcSfk-h7nEA7j2nMhvK-S20BAWZOyE8a2cm0b0gCNW2W4CqoIBOMSw\" width=\"259\" height=\"194\"/></p>\n<p><strong>Primary Introduction</strong> : When the introduced variety is well suited to the new environment, it is released fro commercial cultivation without any alteration in the original genotype.</p>\n<p><strong>Secondary Introduction</strong>: The introduced variety may be subjected to selection to isolate a superior variety. Alternatively, it may be hybridized with local varieties to transfer one or few characters from this variety to the local ones these processes are known as secondary introduction.&nbsp;</p>\n</html>",
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      "body": "<html>\n<p><strong>Farm mechanization</strong> embraces the use of tools, implements and machines for agriculture land development, crop production , harvesting, preparation, storage and on farm processing it include three main power sources.</p>\n<p>&nbsp;1 &nbsp;.Human</p>\n<p>2. Animal</p>\n<p>3. Mechanical</p>\n<p><img src=\"http://www.livemint.com/rf/Image-621x414/LiveMint/Period2/2016/07/08/Photos/Processed/[email protected]\"/></p>\n</html>",
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      "body": "<html>\n<p>&nbsp;&nbsp;<strong>Islam</strong> began in Arabia and was revealed to humanity by the Prophet Muhammad. Those who follow <strong>Islam</strong> are <strong>called</strong> Muslims. Muslims believe that there is only one God. &nbsp;Islam the religion of the Muslims, a monotheistic faith regarded as revealed through Muhammad (SAW)as the Prophet of Allah.</p>\n<p><img src=\"https://dashboard.masjidku.id/images/articles/6e8767e33588f54a3908eb7870219959b88ba562.jpg\" width=\"1280\" height=\"853\"/></p>\n</html>",
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      "body": "<html>\n<p>&nbsp;<strong>Watermelon</strong> is a very good source of vitamin C. It is also a good source of &nbsp;copper, biotin, potassium, &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;vitamin A,vitamin B6 and magnesium. &nbsp;Watermelons are mostly water &nbsp;about 92 percent &nbsp;but this refreshing fruit is soaked with nutrients.&nbsp;</p>\n<p>BENEFITS :</p>\n<p>&nbsp;</p>\n<ul>\n  <li>Helps You Hydrate.</li>\n  <li>Contains Compounds That May Help Prevent Cancer.</li>\n  <li>May Improve Heart Health.</li>\n  <li>May Lower Inflammation and Oxidation Stress.&nbsp;</li>\n  <li>May Help Prevent Oracular Degeneration.&nbsp;</li>\n  <li>May Help Relieve Muscle Soreness.</li>\n  <li>Is Good for Skin and Hair.</li>\n  <li>Can Help Improve Digestion.</li>\n</ul>\n<p><img src=\"https://static.seattletimes.com/wp-content/uploads/2017/07/7b4c85c2-6687-11e7-8665-356bf84600f6-780x520.jpg\" width=\"780\" height=\"520\"/></p>\n<p><br></p>\n</html>",
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