extraction and purification of DNA from plants and plant derived foodstuff and is particularly For example, cetyltrimethylammonium bromide (CTAB) and guanidium thiocyanate (GITC) are often included in protocols for DNA . % Various chemical constituents of DNA extraction buffer allow effective isolation. Chloroform: Mixture of chloroform and isoamyl alcohol (24:1 ratio) equilibrated with a layer of TE buffer (pH 8) on top. Tissue grinding can vary between samples, leading to significant variation in extraction efficiencies and quality of DNA. a vGX. RNA aptamers, specific for different proteins, have been exploited as biorecognition elements to develop specific biosensors (aptasensors). Mix gently to precipitate the nucleic acid. The CTAB procedure would likely work with many gram-negative strains without modification, but gram positives would likely require the addition of lysozyme and proteinase K steps, and may also require modification of medium eg. On with Nicks notes! Wash the resultant DNA pellet with cold alcohol again and centrifuge for retrieval of the pellet. DNA extraction by CsCl density gradient centrifugation: 3. Precipitation: In this final stage, the nucleic acid is precipitated from the solution. 0000001976 00000 n The final precipitate would be eluted and dissolved in the TE buffer. 6R(D$2(>1bD44b9!nGH-NCgVw IIOCebfcDKpC]rY8& "7 k@- Add liquid nitrogen to the tissue and roughly grind the sample into the mortar and pestle. The DNA pellet do not dry and dissolved immediately in 300 l 1xTE, pH 8.0 at 55C for 5-10 minutes. IntechOpen website: https://www.intechopen.com/chapters/49877. PROJECT ON WASTE WATER TREATMENT AND ENERGY PRODUCTION FROM SLUDGE AND SOLID FERMENTATION: TECHNIQUE USED FOR THE PRODUCTION OF FOOD BY MICROBES, Human glucose-6-phosphate isomerase(GPI) or phosphoglucose isomerase(PGI), PHOSPHOLIPIDS, GLYCEROPHOSPHOLIPIDS, SPHINGOMYELIN AND GLYCOLIPIDS, Simulations of Test Reduction Using Pooled Heavy Metals Analysis inCannabis, Quantized space time and Einstein's special theory of relativity.pdf. Water bath set at 65C trailer Sterile Eppendorf tubes and desired reagents. Zymos Quick-DNA Plant/Seed kits use bead beating and column-based purification to provide a simple, rapid workflow for the isolation of inhibitor-free DNA from a variety of plant sources (Figure 1). (CTAB) contained in the extraction buffer. DNA must be purified from cellular material in a manner that prevents degradation. Instant access to millions of ebooks, audiobooks, magazines, podcasts and more. We use this method for extracting genome sequencing quality (i.e. It was adapted from Winnepenninckx B. et al., 1993, TIG: 9 (12), 407 (Technical Tips). Extracted DNA of rose flowers using Cetyl Trimethyl Ammonium Bromide (CTAB) with three different solvents to study the method that gives maximum yields of DNA. Mix vigorously and incubate in water bath at 65C for 1 hr. The lysis of the membranes is accomplished by the detergent The evaluated items included DNA . CTAB extraction buffer. x0pnzmM )3jo.o8e'nn/j!;+RRRJ8F9k^ iN Qfe=\%A9BL YLoa 8C c)o63Z1`*4#q U0 In the CTAB procedure, the first step is breaking down the tissue, and it involves freezing your plant sample using liquid nitrogen. Previous studies demonstrated that Tannin is a proven PCR inhibitor that suppresses PCR amplification and remains bound with the DNA. hb```Lf| Percentages are represented as w/v. 5. Steps in Organic and Inorganic DNA Extraction. DNA extraction is a process of isolating DNA from any biological cell to use it for analysis like PCR, sequencing or restriction digestion. Take 5gms of fresh plant tissue and cut it into small pieces. 13,000 rpm at 4C for 10 min, collect the supernatant to a new eppendorf tube, add 600 L of Now let us come to our major player- CTAB. Cornejo OE, Yee MC, Dominguez V, et al. ii. So these three things (cell wall composition, secondary metabolites and turgor pressure) need advanced treatments. 10% w/v CTAB (Cetyl trimethylammonium bromide), Autoclaved. Then the amount of phenol and chloroform used is greatly reduced (you can use 500 l instead of 5 ml ! From grinding with a mortar and pestle to resuspending sticky DNA pellets, the full CTAB protocol can take approximately two hours to process a small number of samples. DNA extractions using a traditional CTAB-based method with (A) no PVP, (B) 1% PVP and (C) 4% PVP. Take necessary precautions before. This plasmid, pGT5, is a stable multicopy plasmid (2530 copies per chromosome) and can be isolated from cultures at different growth phase and in the temperature range from 75 up to 105oC. Let us see the protocol. For DNA, absorbance at A 260 (also called optical density, OD) is converted into DNA concentration by following method: A 260/OD of 1.0 = a concentration of 50 g/ml of double-stranded DNA . EB (Elution buffer): 5 mM Tris-HCl. extraction method is a simple and non-toxic technique that isolates a high-quality DNA from the whole blood. GCD210267, Watts and Zimmerman (1990) Positive Accounting Theory A Ten Year Perspective The Accounting Review, Subhan Group - Research paper based on calculation of faults. A complete view. Phenol-chloroform and isoamyl alcohol: 4. Bromide (CTAB) protocol (developed by Murray and Thompson in 1980) is appropriate for the 5- Washing . Application of cell and molecular biology techniques, Gene Targeting in Drosophila and Caenorhabditis elegans With Zinc-Finger Nucleases, Mapping chromatin structure in vivo using DNA methyltransferases, Single-molecule and population probing of chromatin structure using DNA methyltransferases, Rapid Detection of Adenovirus from Fecal Specimens, Use of DNA Fingerprinting to Detect Genetic Rearrangements in Human Cancer, Delivery of DNA Vaccines Using Electroporation, In planta expression of oomycete and fungal genes. It is made up of a long phenolic ring and secreted during tissue damage, thus during the lysis process. KIT METHOD OF DNA EXTRACTION. 4. (C++) the answers of assignment ( c++, 2nd edition), Cmo activar Office 2019 gratis y sin programas, Assignment 1. Following plant cell lysis, RNase A is added to digest RNA, and DNA is separated from other cellular components using phenol/chloroform extraction, which separates the sample into two distinct aqueous and organic phases after centrifugation. Weve updated our privacy policy so that we are compliant with changing global privacy regulations and to provide you with insight into the limited ways in which we use your data. conditions, the detergent i. CTAB, which is more stable in alcohol than in water, can be washed Garfinkel AR, Otten M, Crawford S. SNP in Potentially Defunct Tetrahydrocannabinolic Acid Synthase Is a Marker for Cannabigerolic Acid Dominance in Cannabis sativa L. Doyle J, Doyle J. Various chemical constituents of DNA extraction buffer allow effective isolation. Contrary, animal cells dont have turgor pressure, as it is not actually required. The salting-out method is a non-toxic DNA extraction method described by Miller, Dykes, and Polesky in 1988. Do not sell or share my personal information, 1. polysaccharides in a high-salt environment. Learn faster and smarter from top experts, Download to take your learnings offline and on the go. ucdavis. t>V nk+ex!J};0R6B7wRR: %]?i=UXW0l4uRvREu?*js\2cWa>#T|n}%VV?WyuT"8}r6a}s8mP*bL {iWMs(U /M_]{VGZ"\+ Common steps in plant DNA extraction are. Spin at ~10,000 g in microfuge (cold if possible) for 10 min. 4. Under these conditions, proteins, phenolic Start with sample preparation, clean the tissue and prepare it for homogenization. (100 mM Tris-HCl pH 8, 2% (w/v) CTAB, 20 mM EDTA, 1 M NaCl, and 1% (v/v) I will also explain variations and concentrations of different CTAB buffers that you can try and use. )Z$m|h_8"dOtv#P%w;:K&z]x.A`5.AZ%q9;eU! insoluble complex with the polysaccharides. However, other homogenization such as rotor-stator homogenizer or bead mills can be suitable. However, the CTAB-like buffer system can do better with such hard tissues. There are no repetitive and lengthy phase separation steps or hazardous reagents used, so you can further streamline your labs operations and protect the safety of key personnel. Its better to leave a trace of ethanol and water in the pellet (and have it dissolve easily), than to remove all moisture, and then have great difficulty dissolving the pellet. iqy8D!fWJ64. Load 2-4 L of isolated plant genomic DNA in 0% agarose gel and 2- Digestion . This protocol originally came to us from Evelyne Derelle at Observatoire Ocanologique, Banyuls sur Mer. c*f(M=;"]txGyFUHIEW8[WE%:8ioGSTzPF*G@ ioE Such studies stamp CTAB as a major constituent of plant DNA extraction. EB wont give as much protection as TE, which also protects against nucleases. Rl%= For this purpose, the homogenized sample is first treated with the Our binding chemistry, wash solutions, and spin column technology remove polysaccharides, lipids, and other common downstream inhibitors and contaminants providing ultra-pure DNA, with minimal loss. Fast ID contains powerful denaturants and DNA-selective Discard pellet and take supernatant in another tube, Wash the sample with the alcohol (minimum 2 wash). Other methods of DNA extraction include salting out , cesium chloride density gradients , and chelex 100 resin [16, 17]. if(typeof ez_ad_units!='undefined'){ez_ad_units.push([[336,280],'geneticeducation_co_in-leader-2','ezslot_9',168,'0','0'])};__ez_fad_position('div-gpt-ad-geneticeducation_co_in-leader-2-0'); No universal CTAB DNA extraction protocol exists, keep in mind this thing. Precipitation. The extraction is repeated on the aqueous phase until it becomes completely clear, and all DNA is collected. Angeles JGC, Laurena AC, Tecson-Mendoza EM. OPS Diagnostics website: https://opsdiagnostics.com/notes/protocols/ctab_protocol_for_plants.htm. The SlideShare family just got bigger. After pouring the alcohol off the pellet and drying, the DNA can be re-suspended in a buffer such as Tris or TE. Use PCI (phenol: chloroform: isoamyl alcohol) in combination with the CTAB buffer. It was developed in the 1980s and has been used ever since, with various modifications for different plant species.2-5. ORGANIC METHOD OF DNA EXTRACTION 2. n> ; [Content_Types].xml ( ]o '?XV1qvJ~ OV! Grind the tissue in a mortar and pestle and add Urea buffer (which contains Urea, Tris, EDTA and NaCl). National Toxicology Program website: https://ntp.niehs.nih.gov/ntp/roc/content/profiles/chloroform.pdf. [2] CTAB binds to the polysaccharides when the salt concentration is high, thus removing polysaccharides from solution. I already have explained the different types of polysaccharides and polyphenols present in plants and required chemical modifications accordingly. However, care must be taken while handling liquid nitrogen. stream Sterile spatulas INSTRUCTION: If youre working with a challenging plant species, such as cacao and cannabis, try out our state-of-the-art BashingBeads for more complete lysis and more consistent yields (Figure 2).11-14. Add 50 l CTAB, vortex, incubate 60C for 20 min, occasionally mixing by inversion of tube. I .g$!=n]BQ|nTr|0T5t4@XIR$fwyVIszC9Er(ze*RzoI$M%8FM=Mi fR{X[:[F R=XFBdvk>*H7&NIP'B.4GsXUS6j'^p2+#hY[xBYUJuk (L)4/vB`])tS2\YI!MSCKVN",NAN}/DKHFR In addition, working with phenol and chloroform is also a biosafety hazard: Phenol can cause chemical burns and chloroform is a potential carcinogen.6,7 For many food testing labs, the use of these toxic chemicals is a major concern. Leaf samples of two diseased and two asymptomatic plants were collected, and DNA was extracted using a modified CTAB DNA extraction method (Murray & Thomson, 1980). ice-cold ethanol (1000 l) to precipitate DNA incubate at -20C for at least 1 hour. dissolved in the aqueous solution are separated from the lysis homogenate. When the cell membrane is exposed to the inactivation of cellular nucleases and separation of the desired genomic DNA from cellular The DNA was used in nested or single PCR assays to detect the 16S rRNA and SAP genes using the phytoplasma 16S rRNA universal primer pairs P1/P7 and R16F2n/R16R2 (Deng & Hiruki . PVP- PolyVinylPyrrolidone binds with the phenolic ring of tanning and prevents its interaction with DNA. After it becomes homogeneous, take the mixture into the 2ml Eppendorf tube. The instructor can "cheerlead" to encourage ample cells are collected. Dellaporta (1983) and Maroof et al., (1984) have optimized the existing protocols and developed excellent methods for plant DNA extraction. May need to be heated to 50C to dissolve. It actually challenges our knowledge to optimize things in our lab. endstream endobj 38 0 obj <> endobj 39 0 obj <>/Font<>/ProcSet[/PDF/Text]>>/Rotate 0/Type/Page>> endobj 40 0 obj <> endobj 41 0 obj <> endobj 42 0 obj <> endobj 43 0 obj <> endobj 44 0 obj <> endobj 45 0 obj <> endobj 46 0 obj <> endobj 47 0 obj <>stream These two solvents are generally considered hazardous. Additionally, environmental and ethical issues have a higher weight in the consumer's choice to buy honey as a natural product. METHODS OF DNA EXTRACTION Copyright 2023 StudeerSnel B.V., Keizersgracht 424, 1016 GC Amsterdam, KVK: 56829787, BTW: NL852321363B01, Managerial Accounting (Ray Garrison; Eric Noreen; Peter C. Brewer), Principles of Marketing (Philip Kotler; Gary Armstrong; Valerie Trifts; Peggy H. Cunningham), The Importance of Being Earnest (Oscar Wilde), English (Robert Rueda; Tina Saldivar; Lynne Shapiro; Shane Templeton; Houghton Mifflin Company Staff), Mechanics of Materials (Russell C. Hibbeler; S. C. Fan), Auditing and Assurance Services: an Applied Approach (Iris Stuart), Marketing-Management: Mrkte, Marktinformationen und Marktbearbeit (Matthias Sander), Handboek Caribisch Staatsrecht (Arie Bernardus Rijn), Junqueira's Basic Histology (Anthony L. Mescher), Big Data, Data Mining, and Machine Learning (Jared Dean), Frysk Wurdboek: Hnwurdboek Fan'E Fryske Taal ; Mei Dryn Opnommen List Fan Fryske Plaknammen List Fan Fryske Gemeentenammen. A comparison of DNA extraction methods used in research labs as opposed to classroom labs. Looks like youve clipped this slide to already. Take 100 mg tissue, homogenate Summary Aptamers are an alternative to antibodies in their role as biorecognition elements in analytical devices. The successive treatment with 70% ethanol allows an Wash cell pellet in TE (resuspend then pellet again) before lysis if nuclease activity is a problem. 0000002720 00000 n The method is fast,. Remember that genomic DNA means all the DNA, so this method extracts both chromosomes and plasmids. We've updated our privacy policy. PK ! And eventually helps plants to stand straight. 4UYG$(. There are five basic steps of DNA extraction that are consistent across all the possible DNA purification chemistries: 1) disruption of the cellular structure to create a lysate, 2) separation of the soluble DNA from cell debris and other insoluble material, 3) binding the DNA of interest to a purification matrix, 4) washing proteins and other Elution buffer (EB) is just Tris-HCl, which gives the DNA some protection due to its pH buffering. Results indicated that out of five extraction methods, the use of SDS resulted to the highest quality and quantity of DNA, followed by the CTAB method. 3. In this case, after adding TE and NaCl to cell pellet, add lysozyme to 1 mg/ml, and incubate 1 hour at 37C. Be sure you have the proper safety protocols in place before starting your DNA extractions. You choose the most appropriate method ! (BBTV) using PCR RNA Extraction from infected Okra and Cabbage samples from Cameroon, using CTAB method of extraction Show more Show less Education Ladoke Akintola University of Technology . Activate your 30 day free trialto continue reading. By accepting, you agree to the updated privacy policy. Jun 2012 - Dec 2012. Not sterilized. HITS-CLIP (CLIP-Seq) for mouse Piwi proteins. separation of the aqueous and organic phases. Redissolve pellet in 100 l EB. The mantra to success in plant DNA extraction is to grind tissue well, hard, and rough until fine homogenization. 1. Genomic DNA extraction and amplicon generation. As a result, you may leave DNA behind or carryover contaminants from the interphase and organic phase, lowering your overall DNA yield and quality. 0000000893 00000 n Get news, product info, tips, industry updates, events, freebies, and more delivered right to your inbox. For instance, the DNA extraction setup and buffer which works for animal tissue, cant work for plants. Fast ID is able to extract DNA from a wide variety of sample types. Grind 50 mg of plant samples into powder in liquid nitrogen using pre chilled mortar and pestle. To keep up with this rapidly advancing field, DNA extraction protocols must be robust, flexible, consistent, and fast. The CTAB Method: DNA Extraction from Plant Leaves and Seeds To overcome the challenges presented by plant tissues, the cetyltrimethylammonium bromide (CTAB) method has become the "go-to" protocol for DNA extraction and purification from leaves and seeds. Enter the email address you signed up with and we'll email you a reset link. The procedure for fixing the aptamer onto these transducers and for monitoring the interaction with the target protein is shown here. In addition, the presence of turgor pressure, all these things creates problems in plant DNA extraction. "Processing with GMO detection method and confirmation of results for corn, soy, cotton and tomato."-Microorganisms' cultivation.-DNA extraction (CTAB ptotocol) -Method confirmation according to ISO 17025.-PCR (Polymerase chain reaction) . Centrifuge the precipitate for 10,000rpm for 2 minutes and collect the pellet (remove the supernatant). Extraction of high-quality genomic DNA from different plant orders applying a modified CTAB-based method. 0000003202 00000 n UYZsOhq9_&`YWJ+lZ^m]QfPZ^&UTj`F,M^Mmh!.8MLAnJ^gM>LXI=uC2s Ck@1o(T|! Invert tube on paper towel with lid open for ~5 min to drain last bits of 70% ethanol, then transfer to 60C incubator (OK if tube is lying on its side, but lid open) for 10 min to dry residual ethanol. DNA isolation methods are often modified and optimized for different cell types or sample sources. Be careful, take the only supernatant, and dont mix the supernatant with the debris. The method needs to be modified for use on Gram-positives or yeast etc, by adding on extra lysis treatments at the front end of the protocol. Under these There are two reasons for that. Take a sample in the Eppendorf tube and centrifuge it at 12,000rpm for 2 minutes. Results from six grams of leaf tissue finely ground using a mortar and pestle, then aliquoted (1 g) for each extraction. Caution must be taken when working with liquid nitrogen for the first grinding step as it can rapidly freeze skin tissue and cause cold burns even with short exposure. A fragment analyzer Qsep100, that uses a capillary gel electrophoresis system, allowed . CTAB BUFFER 500 ml-1 140mM Sorbitol 12.8 g 220mM Tris, pH 8 55 ml of 2M 22mM EDTA 22 ml of 0.5M 800mM NaCl 80 ml of 5M 1% Sarkosyl 5 g 0.8% CTAB 4 g Combine, check pH = 8, autoclave. If that all seems like a bit much, you arent wrong. ?1tr XE.8TC!qC^t5W3C"w+| 4(XH@CaqB"'1"&)l7%$,Y$DNlNsRF =WPj'Y~u[.qA$D fd9AV'qSFD#>-JBpl W:LlC}P?.EBl!|J)GaABAUoR=$SXjg!K TQQcDxJ$Jh>64=6S[*z~0(OL The cells in a sample are separated from each other, often by a physical means such as grinding or vortexing, and put into a solution containing salt. Clipping is a handy way to collect important slides you want to go back to later. NaCl. 4 0 obj x\KsFW1XD$I~,5>x,}HeVVwmk-m?X/VW|\ 5. uc(j%e\k&5|[CY [,' u8p;le n0%j7giL (0xS6mnf6tW Instead of fuzzy bands, clear and sharp bands can be achieved by using a pinch of CTAB into the SDS PAGE. DNA extraction by chromatography: 2. There are also more than 20 steps in the protocol and as the number of samples increases, the amount of time needed to complete DNA extraction increases substantially. To browse Academia.edu and the wider internet faster and more securely, please take a few seconds toupgrade your browser. Techniques covered include genomic DNA extraction using cetyl trimethylammonium bromide (CTAB) and chloroform extraction, chromatographic techniques, ELISA, hybridization, gel electrophoresis, dot blot analysis and methods for studying polymerase . DNA extraction is the process of isolating DNA from the cells of an organism isolated from a sample, typically a biological sample such as blood, saliva, or tissue . Cell preparation and extraction techniques. Q TP? The aqueous phase is collected, and the recovered DNA is precipitated out with isopropanol. After the cell and organelle PowerPoint: Audio and Video Product Design: From Sketch to CAD . Edwards et al. Free access to premium services like Tuneln, Mubi and more. unsheared) DNA that can be used for large insert libraries. Happily, there are plant-specific DNA isolation kits which provide faster, more consistent, high-purity DNA extraction than the conventional CTAB protocols and variations thereof. (1991) used 0.5% SDS with the CTAB buffer and developed a simple plant DNA extraction protocol. Transfer the supernatant to a new eppendorf tube and add 600 L chloroform, mix and spin at Treat the sample with PCI, centrifuge it, and collect the supernatant part. Select a plant species and prepare the protocol and chemicals accordingly. p|`ylk/?|+.NB/;3a"a~w Y"nX#%C %Ss`2*;2%R=aX&~-o%LZax [C? ?MlB`>8O`-51H>tr$Tt=eyP Le|\! Liquid nitrogen (if available) Under these Nonpolar molecules migrate into the organic phase and leave behind DNA and other polar molecules in the aqueous phase. UV absorbance spectrometry. homogenate to sterile Eppendorf tube. The proteinase K step is additional (you can use it if necessary). After the tissue becomes a powder, add 500 L of CTAB extraction buffer and beta-mercaptoethanol, grind it again followed by vortexing for 5 minutes. INORGANIC METHOD OF DNA EXTRACTION 3. K to 0.5 mg/ml, incubate 60C 1-16 h). By whitelisting SlideShare on your ad-blocker, you are supporting our community of content creators. Carbohydrate, isolation and purification techniques. You can download the paper by clicking the button above. XeQ PRINCIPLE: The extraction of genomic DNA from plant material requires cell lysis, different method, providing an overview before delving deeper into the procedure in a step-by-step approach. In contrast to the original method, the modified CTAB procedure is faster, omits the selective precipitation and CsCl gradient steps, uses less pH 8.0 Autoclaved. Enjoy access to millions of ebooks, audiobooks, magazines, and more from Scribd. step (Lysis of White blood cells, WBC) 3- Phase . Lysis of cell membrane: The first step of the DNA extraction is the rupture of the cell DNA extraction Method for plant sample using CTAB method. Prepare a working solution from stock. Activate your 30 day free trialto unlock unlimited reading. Instant access to millions of ebooks, audiobooks, magazines, podcasts and more. | DNA Extraction Method 2. The CTAB method is biochemically simple, easy to learn, and relatively cheap to perform. Extraction(CTAB) Buffer 1.4 M Na Cl Euphrasia nankotaizanensis (Orobanchaceae) is a rare alpine herb that is endemic to Taiwan. Main Outcome Measures: Successful extraction of DNA Results: PCR, RFLP and DNA sequencing gave the expected results in all the DNA samples extracted by all the three methods (boiling, microwave irradiation and the commercial kit). Use proteinase K overnight at room temperature or for 2 hours at 60C. J w9K|F>.-c#0G{3F)2jSV7\snE>rl gCe:"6zkO( The DNA is insoluble in the alcohol and will come out of solution, and the alcohol serves as a wash to remove the salt previously added. Add 400 l TE and 100 l NaCl (5 M) to pellet, resuspend cells by vortexing. the DNA purity and therefore quality. Centrifuge the sample at 10,000rpm for 2 minutes and take supernatant to another tube. Safe Use of Phenol | Safety Services. We've encountered a problem, please try again. In this study, we demonstrate a modified CTAB method for the isolation of DNA to maximize the quality and yield from small aphids. Phenol, chloroform and isoamyl alcohol DNA extraction. Extraction of DNA, RNA, and protein is the basic method used in molecular biology. The first is a small (3.5 kb) cryptic plasmid found in a high copy number in the marine species P. We've updated our privacy policy. See how we can help you extract high-quality plant DNA, in less time. This method will help the researchers to efficiently isolate DNA from . Procedure 1. aqueous solution with chloroform. Grow culture in 5 ml broth, pellet cells (~3000 g, 10 min), discard supernatant. To make powder with liquid nitrogen or crush it in sterilized pestle mortar with 2 mL of The following stock solutions and reagents were prepared by mixing the standard amount of chemicals. In this article, we will discuss plant DNA extraction and CTAB buffer. Ethanol and NaCl are used to remove plants polysaccharides. 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For example, some plants need only simple grinding in mortar and pestle, while some need liquid nitrogen. However, other chemicals like RNase, proteinase K, SDS and PCI may be used to improve the quality and yield of DNA.