Puregene dna isolation from tails protocol resource. Comparison of different methods for extraction of dna of. Alternatively, it can be safely stored at 20c for later use. Extraction and purification of dna from whole blood and white.
All genetic analysis starts with the collection of dna samples. Dna was quantified via spectrophotometry using a multilabel. Dna is eluted in buffer ae or water, ready for direct addition to pcr or other enzymatic reactions. Jun 29, 2015 enhance your genetics instruction with the jackson laboratorys teaching the genome generation. To determine whether these data are statistically significant p. Protocols for extraction of dna of fungal cells either are very timeconsuming 3 or show poor release of fungal dna 10 compared to methods of extraction of dna of human cells or viruses. Dna genotek oragene og500 kit for collection and prep t2p kit for extraction the prep t2p kit which is based on ethanol precipitation and the pdpr015 whole sample protocol is recommended. Print bookmark share for purification of archivequality dna from human whole blood, bone marrow, buffy coat, buccal cells, body fluids, cultured cells, tissue, mouse tail, yeast, bacteria pdf 367kb english format. Dna genotek dna saliva collection research oragene og500. Table 4 shows the average yield and purity of dna isolated using the three different genomic dna purification procedures described here. The greater the number of white blood cells, the greater the yield.
Use our gels and reagents for quick, accurate separation and analysis of nucleic acids, including the egel system. Methods for extracting genomic dna from whole blood samples. Puregene dna isolation from ears protocol resource. Things to do before starting preheat water baths to 55c for use in step 3b and 65c for use in steps 3a and. Full protocol list below protocol 1 dna extraction part 1.
Dna purification from buffy coat using the gentra puregene blood kit. Determine empirically which protocol works best for your genotyping. The dna molecule is also responsible for heredity, passing on genetic information from parents to child. This kit can be used to extract dna from mammalian blood, tissues, and cultured cells. Enhance your genetics instruction with the jackson laboratorys teaching the genome generation. Dna, deoxyribonucleic acid, is the molecule of life. Dna extraction protocols thermo fisher scientific in. These are available online in convenient and compact pdf format. Preheat water bath to 65c for use step 19 of the procedure. Alternatively, to prevent shearing of high molecular weight dna, omit steps 79 and remove organic solevents and salt. Purification of archivequality dna from nematodes or nematode suspensions using the gentra puregene tissue kit this protocol is designed for purification of dna from 4090 mg 5075.
Our kingfisher purification systems minimize your contact with samples using automated nucleic acid purification for dna, rna, and plasmids. Evaluation of five methods for total dna extraction from. Puregene dna isolation from tails protocol resource labnodes. The approximate yields are 1650 ug dnaml whole blood. Protocol skip steps 16 from the gentra puragene whole blood gdna extraction protocol start at step 7 below. The companys oragenedx and oracollectdx product lines are the first and only fda 510k cleared. Up to 20 ml of blood may be processed in one tube in this protocol. This protocol can be used for extracting dna from blood volumes ranging from 011mls. Dna purification from a buccal brush using the gentra puregene buccal cell kit this protocol is for purification of genomic dna from 1 buccal brush using the gentra puregene buccal cell kit. Dna purification from a buccal brush using the gentra puregene buccal cell kit this protocol is for purification of genomic dna from 1 buccal brush using. A dna extraction protocol for tannin rich eucalyptus and pinus samples which also works very well for acacia, populus, oak and cotton.
Free and easy to use, the open science framework supports the entire research lifecycle. The protocol describes the preliminary harvesting of bacteria and incubation with lysozyme to lyse their cell walls before dna purification. Sep 20, 2016 in the twostep lysis approach used in puregene kits, the first step lyses red blood cells using detergents, such as sodium dodecyl sulfate sds and triton x100. If buffy coat preparation contains red blood cells, treat sample with rbc lysis solution by beginning with step 1 below. Isolate a suitable piece of tissue and place in a uvcrosslinked 1. A simplified universal genomic dna extraction protocol.
Forceps are to be sterilized in fine science tools heat block at. Genomic dna extraction purelink high throughput isolation of pcr products using chargeswitch pcr cleanup iprep genecatcher gdna blood kit for purification of gdna from human blood using the iprep purification instrument. An overview of these kits has been included in table 3. This is a protocol for extracting dna from mouse tails for genotyping. Dna extraction techniques included in table 1 will be. For purification of archivequality dna from human whole blood, bone marrow, buffy coat, buccal cells, body fluids, cultured cells, tissue, mouse tail, yeast, bacteria pdf 367kb.
Protocol puregene dna extraction from fresh frozen tissue. Genomic dna yields from a specific volume of blood will vary depending on the number of nucleated white blood cells present. Grind the tissue into a powder under liquid nitrogen or on an ice bath. Laboratory reagents commonly used for each stage of the nucleic acid extraction protocol are included in this table in order to highlight similarities and differences between them. The dna was purified following protocol 400244 for 4 ml saliva samples collected in oragene kits. Proceed from here with standard protocol starting at step 3 above, copied below. Dna extraction methods from whole blood samples that are generally used in research facilities worldwide.
Genomic dna extraction protocol for pcr dna extraction protocol 1. Comparison of 7 methods for extracting cellfree dna from. Dna purification from tissue using the gentra puregene tissue kit, page 39, if processing 510 mg tissue. The purified dna is free of protein, nucleases, and other. To facilitate a fast but effective dna extraction, fifteen 15 clinical samples were extracted with both a puregene qiagen, hilden, germany and genolyse dna extraction kit hain lifescience gmbh, nehren, germany. High molecular weight dna, which was essential for early rflp methods, may be obtained most effectively with organic extraction. Every living organism has dna in each cell of the organism and each molecule of dna carries the blueprint for that organism. This protocol is designed for purification of total dna from grampositive bacteria. Pdf standard plant dna extraction protocols call for samples of newly.
The lack of consensus regarding which extraction method is better for the efficient capture of such dna might be partially responsible for the large disparities in the literature, which are. Add 20 l dna hydration solution 20 l will give a concentration of 50 ngl if the total yield is 1 g dna. Osur, focuses on providing highquality biological sample collection products and endtoend services for human genomics, microbiome and infectious disease applications. Gentra puregene handbook 062011 5 kit contents plus gentra puregene blood kit 3 ml 30 ml 120 ml ml ml catalog no. Sample processing by our method required approximately 90 min.
Background dna extraction is a routine step in many insect molecular studies. Vortex using light pulses except in step 7 where thorough vortexing is key. Laboratory reagents commonly used for each stage of the nucleic acid extraction protocol. Preheat water bath to 37c for use in step 8 of the procedure. Gentra puregene qiagen dna isolation carl baker, university of washington, 40114 1. Salting out protocol need 10 different reagents for dna extraction, time required is 2 h and 47 min and an overnight step included in processi ng time.
This kit is designed to deliver very high molecular weight dna average size. Protocols for extraction of dna of fungal cells either. Wga with complete genome coverage from single eukaryotic cells or 1 ng genomic dna simple 3step protocol requires only 2 hours and 20 minutes, start to finish consistent yields of up to 2535 g dna from 1 eukaryotic cell. Ribonuclease treatment usually with rnase a then removes. Osf protocol puregene dna extraction from fresh frozen. A quick dirty prep is usually sufficient, while some genotyping may work better with highly purified dna. Comparison of six commercial dna extraction kits for. This protocol can be used for extracting dna from blood volumes ranging from 0. Dna is eluted in buffer ae or water, ready for direct addition to pcr or other enzymatic. Protocols for dna extraction, pcr and gel electrophoresis. For dna extraction, the heat treatment protocol 22 was used. This method is suitable for 96 well arrays and makes use of pvpp or. Frozen blood samples should be thawed quickly in a 37c water bath with mild agitation and stored on ice before beginning this protocol.
Genomic extraction from a few hundred worms will give you a very small dna pellet, which is extremely hard to visualize and easy to loose, so we. Collect reliable samples with an allinone system for the collection, stabilization and transportation of dna from saliva. Show details scalable purification of archivequality dna from 100 to 5 x 108 cultured cells using the gentra puregene cell kit en. Dna purification protocol from 10 ml buffy coat prepared from. This method is suitable for 96 well arrays and makes use of pvpp or pvp 360 to mop up polyphenols. Salting out protocol need 10 different reagents for dna extraction. If possible, tap tube periodically to aid in dispersing the dna. This protocol is designed for purification of dna from 100150 mg dried or 200400 mg fresh leaf tissue using the gentra puregene cell kit. This protocol is for extracting genomic dna from fresh or frozen blood samples. Two samples per patient were pooled together in 700 l of phosphate buffer. Kits available for dna extraction and purification from mammalian cells and tissue are discussed below. Purification requires no phenolchloroform extraction or alcohol precipitation, and involves very little handling. Organic extraction, sometimes referred to as phenol chloroform extraction, has been in use for the longest period of time and for many years was the most widely used method for dna extraction. Do not heat samples except when doing the rnase a step.
Rapid extraction method of mycobacterium ulcerans dna from. In salting out technique of dna extraction, high salt concentration used with phenol and chloroform hazardous organic solvent for protein precipitation. Dna purification from compromised blood samples using the gentra puregene blood kit this protocol is for purification of genomic dna from 10 ml compromised whole blood using the gentra puregene blood kit. Incubate oragene saliva samples at 50c in a water incubator for a minimum of 1 hour or in an. Add 50 l dna hydration solution 50 l will give a concentration of 500 gml if the total yield is 25 g dna.
The approximate yields are 1650 ug dna ml whole blood. Extraction and purification of dna from whole blood and. Then, a second step ensures lysis of white blood cells to release the cell nucleus, genomic dna, and rna figure 1a. These results suggest that the ns kit offers the highest degree of cmv dna recovery with the broad range of specimen types evaluated. Small colonies were transferred into 2 ml eppendorf tubes with 1. Optimum yields of dna are achieved when the blood samples are processed within five days of being obtained from the patient. For long term storage it is convenient to leave the dna in the presence of ethanol.
To prevent degradation of dna, avoid vigorous vortexing and heating. Pdf dna extraction protocols from dormant buds of twelve. Extracting the entire oragene sample allows for maximum na recovery and concentration. This protocol is for purification of genomic dna from 10 ml compromised whole blood using the gentra puregene blood kit. Need cells because thats where dna is a in the nucleus. Dna purification from compromised blood samples using the gentra puregene blood kit this protocol is for purification of genomic dna from 10 ml. Why do you need dna genetic testing, body identification, analysis of forensics evidence dna needs to be purified away from proteins and other cellular contaminents. A variety of methods have been used to isolate dna molecules from insects, and many commercial kits are available. Dna purification kit dna purification protocol from 10 ml buffy coat prepared from 10 ml whole blood 50 ml tube prep expected yield range 200400 g dna cell lysis thaw buffy coat for 12 minutes at 37c. Methods for extracting genomic dna from whole blood. To facilitate a fast but effective dna extraction, fifteen 15 clinical samples were extracted with both a puregene qiagen, hilden, germany and.
To determine whether these data are statistically significant p 0. Dna extraction from insects by using different techniques. Dna genotek oragene og500 kit for collection and prep t2p kit for extraction the prep t2p kit which is based on ethanol precipitation and the pdpr015 whole sample protocol is recommended for extraction of dna from saliva samples. Phenolchloroform extraction an overview sciencedirect topics. For all samples, dna was extracted using a modified dna extraction protocol puregene. Intended use the fiberprep dna extraction kit is intended for the extraction of dna from fresh blood samples or cultured cells for molecular combing applications.
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