DNA IQ System-Small Sample Casework Protocol Technical Bulletin
Contents
1. Mix liquid sample prepared Lysis Buffer and resin Vortex and incubate at room temperature for 5 minutes Vortex and place in magnetic stand Carefully discard solution without disturbing resin Wash with prepared Lysis Buffer Wash three times with 1X Wash Buffer Air dry at room temperature Add Elution Buffer and incubate at 65 C for 5 minutes Remove tubes from heat vortex and place in magnetic stand Remove DNA solution 3238MB Figure 2 Schematic of DNA purification from liquid samples using the DNA IQ System See Section 4 C for a detailed protocol Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Part TB296 Printed in USA Page 4 Revised 12 10 Promega 3 Sample Types Examined DNA from the following sample types have been successfully purified at Promega or by external forensic laboratories Due to the nature of casework samples i e the samples may have been exposed to environmental factors for long periods of time and the amount of biological material may be limiting DNA yields may vary and DNA may not be obtained from all samples Please see the most updated list at www promega com dnaiqsamples Tissue masses including hair bone and sperm require a proteinase K digestion to obtain reliable amounts of DNA Contact Promega Technical Services genetic p2. DC6500 PowerPlex 1 2 System 100 reactions DC6101 Not For Medical Diagnostic Use U S Pat Nos 6 027 945 6 368 800 and 6 673 631 Australian Pat No 732756 European Pat Nos 0 895 546 and 1 204 741 and Mexican Pat No 209436 have been issued to Promega Corporation for methods of isolating biological target materials using silica magnetic particles and simultaneous isolation and quantitation of DNA Other patents are pending 2010 Promega Corporation All Rights Reserved AluQuant MagneSphere PolyATtract and PowerPlex are registered trademarks of Promega Corporation DNA IQ is a trademark of Promega Corporation ART is a registered trademark of Molecular Bio Products Inc Biomek is a registered trademark of Beckman Coulter Inc Chelex is a registered trademark of Bio Rad Laboratories Inc Freedom EVO is a registered trademark of Tecan AG Corporation FTA is a registered trademark of Flinders Technologies Pty Ltd and is licensed to Whatman Products may be covered by pending or issued patents or may have certain limitations Please visit our Web site for more information All prices and specifications are subject to change without prior notice Product claims are subject to change Please contact Promega Technical Services or access the Promega online catalog for the most up to date information on Promega products Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 P
3. Do not dry for more than 20 minutes as this may inhibit removal of DNA Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Part TB296 Printed in USA Page 10 Revised 12 10 Promega 13 Add 25 1001 of Elution Buffer depending on how much biological material was used A lower elution volume ensures a higher final concentration of DNA 14 Close the lid and vortex tube for 2 seconds at high speed Incubate at 65 C for 5 minutes 15 Remove the tube from the heat source and vortex for 2 seconds at high speed Immediately place on the magnetic stand Tubes must remain hot until placed in the magnetic stand or yield will decrease 16 Carefully transfer the DNA containing solution to a container of choice Note DNA can be stored at 4 C for short term storage or at 20 or 70 C for long term storage 4 D Processing of Sperm Containing Samples The following approach describes DNA purification from differentially extracted samples of sperm and nonsperm fractions using the DNA IQ System This approach has been successfully performed in a forensic laboratory setting 1 The sample of interest is first processed using the laboratory s validated differential extraction protocol 2 including the extraction of biological material from the solid support proteinase K digestion in the absence of DTT and pelleting and
4. microcentrifuge tube 3 Vortex the resin Lysis Buffer mixture for 3 seconds at high speed to ensure suspension of resin and add 100ul of the mixture to the tube containing the liquid sample The resin Lysis Buffer mixture should be mixed again if the resin begins to settle while dispensing aliquots 4 Vortex the sample Lysis Buffer resin mix for 3 seconds at high speed Incubate 5 minutes at room temperature Vortex mixture for 3 seconds once every minute during this 5 minute incubation 5 Vortex for 2 seconds at high speed Place tube in the magnetic stand Separation will occur instantly Note If resin does not form a distinct pellet on the side of the tube vortex the tube and quickly place it back in the stand 6 Carefully remove and discard all of the solution without disturbing the resin pellet on the side of the tube 7 Add 100ul of prepared Lysis Buffer Remove tube from the magnetic stand and vortex for 2 seconds at high speed 8 Return tube to the magnetic stand and remove and discard all Lysis Buffer 9 Add 100ul of prepared 1X Wash Buffer Remove tube from the magnetic stand and vortex for 2 seconds at high speed 10 Return tube to the magnetic stand Dispose of all Wash Buffer 11 Repeat Steps 9 and 10 two more times for a total of three washes Be sure all of the solution has been removed after the last wash 12 With the tube in the magnetic stand and the lid open air dry the resin for 5 minutes O
5. washing of the sperm cells The following steps are to be followed once the sperm and nonsperm fractions have been separated There is no need to digest the sperm pellet with proteinase K and DTT as the prepared Lysis Buffer in Step 1 will effectively disrupt the sperm cells after the initial proteinase K digestion Increasing the DTT concentration in the prepared Lysis Buffer to 60mM can improve the recovery of DNA from the sperm fraction To prepare Lysis Buffer with a final concentration of 60mM DTT add 6ul of 1M DTT to 100ul of DNA IQ Lysis Buffer instead of 141 of 1M DTT as described in Section 4 A 1 Add at least 2 volumes minimum 100ul of prepared Lysis Buffer and 7 1 of resin to the sperm pellet 2 Vortex for 3 seconds at high speed and incubate at room temperature for 5 minutes 3 Proceed to Steps 5 16 of Section 4 C to purify DNA from this sperm fraction 4 Add two volumes of prepared Lysis Buffer and 7ul of resin to the nonsperm fraction Note If desired 100u1 of a 500u1 extraction can be processed This amount typically gives sufficient DNA for analysis Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Printed in USA Part TB296 Revised 12 10 Page 11 Promega 5 Vortex for 3 seconds at high speed and incubate at room temperature for 5 minutes 6 Proceed to Steps 5 16 of Section 4 C t
6. 6 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Part TB296 Printed in USA Page 8 Revised 12 10 Promega 7 Vortex tube for 2 seconds at high speed Place tube in the magnetic stand Separation will occur instantly Note If resin does not form a distinct pellet on the side of the tube vortex the tube and quickly place back in the stand 8 Carefully remove and discard all of the solution without disturbing the resin pellet on the side of the tube Note If some resin is drawn up in tip gently expel resin back into tube to allow re separation 9 Add 100ul of prepared Lysis Buffer Remove the tube from the magnetic stand and vortex for 2 seconds at high speed 10 Return tube to the magnetic stand and discard all Lysis Buffer 11 Add 100ul of prepared 1X Wash Buffer Remove tube from the magnetic stand and vortex for 2 seconds at high speed 12 Return tube to the magnetic stand and discard all Wash Buffer 13 Repeat Steps 11 and 12 two more times for a total of three washes Be sure that all of the solution has been removed after the last wash 14 With the tube in the magnetic stand and the lid open air dry the resin for 5 minutes Q Do not dry for more than 20 minutes as this may inhibit removal of DNA 15 Add 25 1001 of Elution Buffer depending on how much biological material was used A lower elution volume ensures a higher final concentration of DNA 16 Close the lid and vortex the
7. DNA IQ System 100 samples DC6701 This system includes 0 9m Resin 40ml Lysis Buffer 30ml 2X Wash Buffer 15ml Elution Buffer Storage Conditions Store all DNA IQ System reagents at 22 25 C Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Part TB296 Printed in USA Page 2 Revised 12 10 Promega Cut or punch out sample and place in a microcentrifuge tube Add prepared Lysis Buffer and incubate at 70 C for 30 minutes Transfer Lysis Buffer and sample into DNA IQ Spin Basket Centrifuge Remove spin basket and add resuspended resin Vortex and incubate at room temperature Vortex and place in magnetic stand Carefully discard solution without disturbing resin Wash with prepared Lysis Buffer Wash three times with 1X Wash Buffer Air dry at room temperature Add Elution Buffer and incubate at 65 C for 5 minutes Remove tubes from heat vortex and place on magnetic stand Remove DNA solution 3237MB Figure 1 Schematic of DNA isolation from stains on solid material using the DNA IQ System See Section 4 B for a detailed protocol Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Printed in USA Part TB296 Revised 12 10 Page 3
8. Technical Bulletin DNA IQ System Small Sample Casework Protocol INSTRUCTIONS FOR USE OF PRODUCTS DC6700 AND DC6701 PRINTED IN USA Revised 12 10 Part TB296 Promega DNA IQ System Small Sample Casework Protocol All technical literature is available on the Internet at www promega com tbs Please visit the web site to verify that you are using the most current version of this Technical Bulletin Please contact Promega Technical Services if you have questions on use of this system E mail genetic promega com 1 DCSE UVM osier ee a 1 2 Product Components and Storage Conditions eeesesteeeeeeeeeenen 2 3 Sample Types Examined 0 0 ceesesssseseeseeseseeseeseseeseeneeeeseeseeeeaeeneateneeeeeenee 5 4 Protocol for the DNA IQ System esesseeececeeseeeecteneseateneeeeetenee 6 As Preparation Ol Rea Cet is soseri onna EEE i B DNA Isolation from Stains on Solid Material teeeseseeeseeeeeeeeeeeeeen 8 CDNA Isolation Mont Lig uid Samples iceccacsaseacnrinssventareanaiaamnannsein 10 D Processing of Sperm Containing Samples cesses 11 5 Troubleshooting ocscesccpccresecesteoetstcateraersueaensteeroreneceercnens enrmarscomuaanaesaes 12 6 References sssenssssesssesesssesesssessserssssetessterossrerossrteosesreessertessteresssetessteeessreress 13 7 Composition of Buffers and Solutions ceccesesseseeeseeseeeeseeeeneeeens 13 6 Related Products ixiscnsctaexstdayeosysctcranesesaeserenpsnesansnertaave
9. can be stored at room temperature for up to a month if sealed Note If prepared Lysis Buffer forms a precipitate warm solution to 37 60 C Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Printed in USA Part TB296 Revised 12 10 Page 7 Promega 4 B DNA Isolation from Stains on Solid Material The maximum DNA yield will depend on the sample type even samples that contain DNA amounts in excess of the DNA binding capacity of the resin As expected for samples that do not contain DNA amounts exceeding the DNA binding capacity the yield will vary with the sample type and amount Samples containing small amounts of DNA will have high effiencies of recovery as the DNA content approaches the maximum DNA binding capacity efficiency decreases see Figure 1 of the DNA IQ System Database Protocol TB297 1 Place sample in a 1 5ml microcentrifuge tube e g Microtubes 1 5ml Cat V1231 2 Add the appropriate volume of prepared Lysis Buffer Different samples require different volumes of prepared Lysis Buffer see Column 2 of Table 3 for the appropriate volume to add at this point Additional prepared Lysis Buffer may be used to cover entire sample Close the lid and incubate tube at 70 C for 30 minutes Exceptions e Heat sensitive fabrics e g polyester and nylon Extract without heating Leath
10. er Lysis Buffer extraction with or without heat may not work on some leathers Extract in a small volume of aqueous buffer 100 200u1 then add 2 volumes of Lysis Buffer after removing matrix Note For small stains an alternative approach is to place the stained material ina DNA IQ Spin Basket Cat V1221 seated in a 1 5ml Microtube Cat V1231 Add 100 150u1 of prepared Lysis Buffer to the basket Carefully close the lid and incubate at 70 C for 30 minutes Most of the buffer should remain in the basket if the indicated tubes and spin baskets are used Proceed to Step 4 3 Remove the tube from the heat source and transfer the prepared Lysis Buffer and sample to a DNA IQ Spin Basket seated in a 1 5ml Microtube 4 Centrifuge at room temperature for 2 minutes at maximum speed in a microcentrifuge Remove the spin basket Note It is important to centrifuge the prepared Lysis Buffer and stained matrix to obtain maximum recovery 5 Vortex the stock resin bottle for 10 seconds at high speed or until resin is thoroughly mixed Add 7ul of DNA IQ Resin to the sample Keep the resin resuspended while dispensing to obtain uniform results 6 Vortex sample Lysis Buffer resin mixture for 3 seconds at high speed Incubate at room temperature for 5 minutes Vortex mixture for 3 seconds once every minute during this 5 minute incubation Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 35
11. hone 608 274 4330 Fax 608 277 2516 www promega com Part TB296 Printed in USA Page 14 Revised 4 06
12. les add 15ml of 95 100 ethanol and 15ml of isopropyl alcohol to the 2X Wash Buffer For DC6700 400 samples add 35ml of 95 100 ethanol and 35ml of isopropyl alcohol to the 2X Wash Buffer 2 Replace cap and mix by inverting several times 3 Mark label to record the addition of alcohols Label bottle as 1X Wash Buffer Solution can be stored at room temperature Be sure bottle is closed tightly to prevent evaporation Preparation of Lysis Buffer 1 Determine the total volume of prepared Lysis Buffer to be used Table 3 and add 1pl of 1M DTT for every 100ul of Lysis Buffer Note Increasing the DTT concentration in the prepared Lysis Buffer to 60mM can improve the recovery of DNA from the sperm fraction of sperm containing samples Section 4 D To prepare Lysis Buffer with a final concentration of 60mM DTT add 6ul of 1M DTT to 100ul of Lysis Buffer Table 3 Volume of Prepared Lysis Buffer Required Per Sample Sample Lysis Buffer Lysis Buffer Total Volume Liquid samples up to 40ul 100ul 100ul 200u1 1 Cotton swab 250ul 1001 350ul 1 4th CEP swab 250ul 100ul 350ul 1 2 4mm punches of S amp S 903 paper 150ul 100l 250ul 1 3 2mm punches of FTA paper 150ul 1001 250ul Cloth up to 25mm 150ul 1001 250ul 1For use in Section 4 B Step 2 or Section 4 C Step 1 For use in Section 4 B Step 9 or Section 4 C Step 7 2 Mix by inverting several times 3 Mark and date label to record the addition of DTT This solution
13. mock sexual assault samples using the Biomek 2000 and the DNA IQ System Profiles in DNA 5 3 5 2 Gill P et al 1985 Forensic application of DNA fingerprints Nature 318 577 9 7 Composition of Buffers and Solutions Elution Buffer Bone Incubation Buffer 10mM Tris pH 8 0 10mM Tris pH 8 0 0 1mM EDTA 100mM NaCl 50mM EDTA 0 5 SDS Promega Corporation 2800 Woods Hollow Road Toll Free in USA 800 356 9526 Phone 608 274 4330 Printed in USA Revised 12 10 Madison WI 53711 5399 USA Fax 608 277 2516 WWwW promega com Part TB296 Page 13 Promega 8 Related Products Product Size Cat MagneSphere Technology Magnetic Separation Stand two position 1 5ml Z59932 MagneSphere Technology Magnetic Separation Stand twelve position 1 5ml 25342 PolyATtract System 1000 Magnetic Separation Stand 1 each Z5410 DNA IQ Spin Baskets 1 000 pk V1221 Microtubes 1 5ml 1 000 pk V1231 ART 20P Pipet Tip 20 1 960 pk DY1071 ART 200 Pipet Tip 200u1 960 pk DY1121 ART 1000E Pipet Tip 1 000u1 800 pk DY1131 Slicprep 96 Device 10 pack V1391 AluQuant Human DNA Quantitation System 80 determinations DC1010 400 determinations DC1011 Tissue and Hair Extraction Kit for use with DNA IQ 100 reactions DC6740 DTT Molecular Grade Dry Powder 5g V3151 25g V3155 PowerPlex 16 System 100 reactions DC6531 400 reactions DC6530 PowerPlex 1 1 and 2 1 Systems 100 reactions DC6501 400 reactions
14. o purify DNA from this nonsperm fraction 5 Troubleshooting For questions not addressed here please contact your local Promega Branch Office or distributor Contact information available at www promega com E mail genetic promega com Symptoms Causes and Comments Poor yield Too much sample was used Excessive amounts of sample can reduce the efficiency of DNA binding to the resin Use less sample or more resin Poor extraction After heating stain in prepared Lysis Buffer centrifuge buffer with matrix Be sure enough liquid is present to wash out the DNA Excessive drying of resin Do not dry samples for more than 20 minutes as overdrying the resin inhibits DNA elution For sperm containing samples increasing the DTT concentration in the prepared Lysis Buffer to 60mM may improve yield see Section 4 D Poor resin pellet formed The resin settled before the tube was placed in the magnetic stand Samples should be placed in the magnetic stand immediately after mixing Repeat mixing and place tube in stand Excessive input material was used relative to the recommended volumes of reagents Use less initial sample Consult protocols for recommended quantities of initial sample Alternatively use more resin per isolation A proportional increase in resin will allow DNA capture from more initial sample The increase in yield will be roughly proportional to the increase in resin Coloration in final wash or eluted In
15. romega com for the latest information on available protocols Table 1 Types of Samples From Which DNA Has Been Successfully Isolated Using the DNA IQ System Sample Type Promega External Comments Fresh blood Yes Yes Works with the following anti clotting reagents EDTA citrate heparin ACD Frozen blood Yes Yes Old blood may produce lower yields Bloodstains S amp S 903 paper Yes Yes FTA paper Yes Yes Cotton Yes Yes Blue denim Yes Yes Black denim Yes Yes Soil Yes Leather Yes Yes Surface to swab Yes Buccal swabs Cotton Yes Yes Rayon Nes CEP paper Yes Swab to FTA paper Yes Foam swab to paper Yes Cigarette butt Yes Yes Use paper wrapping filter may form gel if heated with prepared Lysis Buffer Toothbrush Yes Soak bristles in prepared Lysis Buffer at 60 C for 30 minutes Envelope Yes Soak in 0 5 SDS before adding 2 volumes of prepared Lysis Buffer Urine Yes Sample from bladder cancer patient Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Printed in USA Part TB296 Revised 12 10 Page 5 Promega Table 2 Samples Requiring a Proteinase K Digestion Prior to Addition of Twice the Recommended Volume of Lysis Buffer Sample Type Promega External Comments Tissue Fresh Yes Yes See the Tissue and Hair Extraction Kit for use with DNA IQ Technical Bulletin TB307 Formalin fixed Yes Ye
16. s See the Tissue and Hair Extraction Kit for use with DNA IQ Technical Bulletin TB307 Hair Yes Yes See the Tissue and Hair Extraction Kit for use with DNA IQ Technical Bulletin TB307 Bone Yes From pulverized bone samples Antler Yes From drill shavings Differential extractions Yes See Section 4 D Requires the Bone Incubation Buffer containing 1mg ml proteinase K for DNA purification Please contact Technical Services genetic promega com for a protocol for DNA isolation from bone samples 4 Protocol for the DNA IQ System Materials to Be Supplied by the User e 95 100 ethanol e isopropyl alcohol e IMDITT e 65 C heat block water bath or thermal cycler e 70 C heat block water bath or thermal cycler for stain or swab extraction e vortex mixer e Microtubes 1 5ml Cat V1231 e DNA IQ Spin Baskets Cat V1221 e aerosol resistant pipette tips e MagneSphere Technology Magnetic Separation Stand twelve position Cat Z5342 e proteinase K to process samples listed in Table 2 Use of gloves and aerosol resistant pipette tips is highly recommended to prevent cross contamination Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Part TB296 Printed in USA Page 6 Revised 12 10 Promega 4 A Preparation of Reagents Preparation of 1X Wash Buffer 1 For DC6701 100 samp
17. sufficient washing Remove all fluid during solution may affect results of washes Be sure that resin is completely downstream assays resuspended during each wash step Be sure a distinct resin pellet is formed during all washes Use less initial sample Perform additional washes with 1X Wash Buffer Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Part TB296 Printed in USA Page 12 Revised 12 10 Promega 5 Troubleshooting continued Symptoms Causes and Comments Resin present in final eluted solution may affect results of downstream assays Resin is occasionally transferred by rapid pipetting or is caught in the meniscus of the final eluate Vortex or mix solution place in the magnetic stand and transfer eluate to new tube Inconsistent amounts of resin Vortex resin stock just before making aliquots Be sure to vortex the resin Lysis Buffer mixture while dispensing aliquots Inconsistent yield may affect results from downstream assays Excessive drying of resin Do not dry samples for more than 20 minutes as overdrying the resin inhibits DNA elution Contaminating nonhuman DNA in the initial sample can decrease yield of human DNA The DNA IQ System captures total DNA including single and double stranded DNA 6 References 1 Greenspoon S and Ban J 2002 Robotic extraction of
18. tube for 2 seconds at high speed Incubate the tube at 65 C for 5 minutes 17 Remove the tube from the heat source and vortex for 2 seconds at high speed Immediately place the tube on the magnetic stand Tubes must remain hot until placed in the magnetic stand or yield will decrease 18 Carefully transfer the DNA containing solution to a container of choice Note DNA can be stored at 4 C for short term storage or at 20 or 70 C for long term storage Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Printed in USA Part TB296 Revised 12 10 Page 9 Promega 4 C DNA Isolation from Liquid Samples This protocol is recommended for liquid samples excluding liquid blood A protocol for DNA purification from liquid blood can be found in the DNA IQ System Database Protocol 1B297 1 Prepare a stock solution of resin and Lysis Buffer by using the ratio of 7 of resin to 93ul of prepared Lysis Buffer per sample prepare extra to allow for losses during pipetting The following equation will help determine the exact volumes to be made Vortex the resin container for 10 seconds at high speed or until resin is thoroughly mixed Number of samples 1 x Zul ul of resin Number of samples 1 x 93 ul ul of prepared Lysis Buffer 2 Mix liquid sample gently and place an aliquot of up to 40ul into a 1 5ml
19. used with a filtration system but tend to give lower yields and require extensive washing to remove the guanidine based lysis buffer Currently available silica magnetic particles tend to give higher yield but also need extensive washing The DNA IQ System uses a novel paramagnetic resin for DNA isolation Using the DNA IQ System to process small casework samples requires two steps For biological material on solid supports the first step provides an easy rapid efficient and almost universal stain extraction method This step is unnecessary for liquid samples The second step uses the paramagnetic resin to purify DNA without requiring extensive washing to remove the lysis reagent This system is designed to rapidly purify small quantities of DNA and give consistent yields for a specific sample type The DNA IQ System has been automated using the Beckman Coulter Biomek 2000 and 3000 Laboratory Automation Workstations and Tecan Freedom EVO 100 liquid handler For more information see the DNA IQ System product profile at www promega com applications hmnid productprofiles automation For more information about implementing these methods contact Promega Technical Services genetic promega com 2 Product Components and Storage Conditions Product Size Cat DNA IQ System 400 samples DC6700 This system includes 3ml Resin 150ml Lysis Buffer 70ml 2X Wash Buffer 50m Elution Buffer Product Size Cat
20. wenmnaseenrinmaelatadene 14 1 Description DNA analysis is playing an increasingly larger role in identifying both humans and animals With the advent of large multiplexes such as the PowerPlex 16 System the amplification and analysis steps have been streamlined However the purification of DNA from a variety of samples is still a rate limiting step in obtaining useful genotypes Several procedures are currently used to purify DNA from samples adsorbed to surfaces Biological material must first be removed from these surfaces This is typically done by soaking the material which may result in inefficient recovery of small samples The DNA must then be purified from inhibitors of PCR amplification and other components that may interfere with accurate quantitation methods Purification methods commonly used such as phenol chloroform extraction use hazardous organic chemicals require multiple centrifugations may result in significant loss of material and can introduce amplification inhibitors Chelex extraction is rapid but frequently leaves amplification inhibitors Purification Promega Corporation 2800 Woods Hollow Road Madison WI 53711 5399 USA Toll Free in USA 800 356 9526 Phone 608 274 4330 Fax 608 277 2516 www promega com Printed in USA Part TB296 Revised 12 10 Page 1 Promega with silica matrices uses the affinity of DNA for silica and does not require organic components Silica filters are convenient when
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