FastDNA® Green SPIN Kit
Contents
1. with a new clean tube 11 Elute DNA by gently resuspending Binding Matrix above the SPIN filter in 100 ul of DES Incubate for 5 minutes at 55 C in a heat block or water bath 12 Centrifuge at 14 000 x g for 1 minute to bring eluted DNA into the clean catch tube Discard the SPIN filter DNA is now ready for downstream applications Store at 20 C for extended periods or 4 C until use www mpbio com 6 Example Data DNA Isolation from Animal and Plant Samples and Gel Electrophoresis DNA from plant samples extracted with the FastDNA Kit Approximately 1 ug of solated DNA was loaded on a 1 2 agarose gel 0 5X TAE Lane 1 0 16g apple stem Lane 2 0 459 red bell pepper seeds Lane 3 0 45g pelargonium root Lane 4 0 45g mature peace lily leaf Lane 5 0 45g ice plant leaf Lane 6 Lambda Hind Ill marker DNA from animal samples extracted with the FastDNA Kit Approximately 1 ug of isolated DNA was loaded on a 1 2 agarose gel 0 5X TAE Lane 1 0 4g rat liver Lane 2 0 5g mouse brain Lane 3 0 45g chicken bone Lane 4 Lambda Hind Ill marker www mpbio com 9 FastDNA Green Spin Kit 7 Table of Typical FastPrep Settings l FastPrep FastPrep HUMAN AND ANIMAL Humn Lysing Matrix D 40sec Sec Thyroid Mose J0mg mg Lysing Matrix D Matrix D 4x 4x 30sec Sec clim 1 1s ue Cw me m TT en sm Cus prm pm TO 00 ET Lum ATC som TT 00 one Pause2. Instruction Manual E FastDNA Green SPIN Kit Rapid Isolation of Genomic DNA from Plant and Animal Tissues Using the FastPrep System gt One Call a 100 P One Source T Storage 7 A World of gt Ambient temperature 15 30 C B I otech n ol Og y Catalog 6540 800 Reagents j Revision 6540 800 10JUL gt A eee www mpbio com MP Biomedicals e 29525 Fountain Parkway e Solon OH 44139 e tel 1 800 854 0530 e fax 1 800 334 6999 FastDNA Green Spin Kit Lyse BIG and FAST with Interchangeable Adapters QuickPrep Adapter TeenPrep Adapter 24 x 2 ml samples 12 x 15ml samples Ll y m nan HighPrep Adapter BigPrep Adapter 48 x 2ml samples 2 X 50 ml samples E 7 2 The FastPrep 24 instrument is delivered with the QuickPrep Adapter www mpbio com FastDNA Green SPIN Kit Rapid Isolation of Genomic DNA from Plant and Animal Tissues Using the FastPrep System 100 Preps storage Ambient temperature 15 30 C Catalog 6540 800 Revision 6540 800 10JUL www mpbio com FastDNA Green Spin Kit TABLE OF CONTENTS 1 Introduction to the FastDNA Green SPIN Kit and the FastPrep Instruments 2 Kit Components and User Supplied Materials 2 1 FastDNA SPIN Green Kit Components 2 2 User Supplied Materials 3 Important Co
3. ach www mpbio com 5 FastDNA Green Spin Kit 2 2 User Supplied Materials FastPrep Instrument see Section 10 Microcentrifuge that can freely spin 2 0 ml tubes Microcentrifuge tubes 2 0 ml and 1 5 ml Rotator or low speed vortex 3 Important Considerations Before Use 3 1 Preparation of SEWS M Wash Solution The FastDNA Green SPIN Kit contains a bottle with 12 ml of Concentrated SEWS M Wash Solution Before using this solution add 100 ml of 100 ethanol and mark on the bottle label the date ethanol was added Ensure that the bottle is securely closed to prevent evaporation and store at room temperature 3 2 Precipitation in CLS TC Buffer If the FastDNA Green SPIN Kit was shipped or stored at a low temperature a harmless precipitate may form in the CLS TC Buffer If a precipitate is seen incubate the bottle in a 45 55 C water bath for several minutes and mix to bring the precipitate back into solution Allow solution to cool to room temperature 3 3 Sample Lysis with the FastPrep Instrument The fill volume in the lysing matrix tube after the addition of the Cell Lysis Solution to the sample should allow sufficient air space in the sample tube for efficient FastPrep Instrument processing MP Biomedicals recommends using 100 200 mg of starting material as long as there is between 250 500 ul of empty space in the tube Sample loss or tube failure may result from overfilling the matrix tube The matrix tube caps mu
4. es To optimize DNA recovery from extremely dry samples leave the lysed sample at room temperature in the Lysing Matrix D tube for an incubation period of 15 minutes to 2 hours after processing in the FastPrep Instrument 3 5 Co Purification of RNA Some tissues i e liver kidney contain very high levels of RNA which may co purify with the genomic DNA If absolute control of RNA contamination is necessary the final eluted DNA can be treated with RNase as per the manufacturer s protocol 4 Safety Precautions Binding Matrix contains components that when in contact with human tissue may cause irritation Wear personal protective equipment to prevent contact with the skin or mucus membranes gloves lab coat and eye protection Consult the enclosed Material Safety Data Sheet for additional details 5 Protocol 1 Add sample to Lysing Matrix D tube Place up to 100 200 mg tissue fresh frozen dried etc or 200 ul of cells suspended in water or isotonic saline solution For blood lymph or tissue culture cells grown in suspension Centrifuge a sufficient volume of culture to provide a pellet size of 50 100 mg wet weight or up to 107 mammalian or plant cells Resuspend pellets in water or isotonic saline to give a maximum suspensionvolume of 200 ul NOTE See section 3 3 for other important guidelines www mpbio com 7 2 9 FastDNA Green Spin Kit Add appropriate Cell Lysis Solution CLS according to table be
5. low Plant tissues 800 ul CLS VF and 200 ul PPS Human and animal tissues cultured cells 1 0 ml CLS TC Homogenize in the FastPrep Instrument for 40 seconds at a speed setting of 6 0 Centrifuge at 14 000 x g for 5 10 minutes to pellet debris Transfer supernatant 700 800 ul to a 2 0 ml microcentrifuge tube and add an equal volume of Binding Matrix Invert to mix NOTE It is important to use a tube that is large enough to allow room for complete mixing of the entire volume during the course of the next step Tubes with conical bottoms are not recommended A 2 0 ml microcentrifuge tube works well at this Step Incubate with gentle agitation for 5 minutes at room temperature on a rotator NOTE A low speed vortex may be used at this point but care must be taken not to shear the DNA Transfer half approximately 800 ul of the suspension to a SPINTM Filter and centrifuge at 14 000 x g for 1 minute Empty the catch tube and add the remaining suspension to the SPIN Filter and centrifuge as before Empty the catch tube again Add 500 ul prepared SEWS M and gently resuspend the pellet using the force of the liquid from the pipet tip NOTE Ensure that ethanol has been added to the Concentrated SEWS M See section 3 1 Centrifuge at 14 000 x g for 1 minute Discard contents of Catch Tube and replace 10 Without any addition of liquid centrifuge a second time at 14 000 x g for 2 minute and replace the Catch Tube
6. m Instruction Manual FastDNA Green SPIN Kit Protocol Revision 6540 800 10JUL 2 Worldwide Ordering and Technical Support United States of America France Serbia Worldwide Headquarters MP Biomedicals France MP Global d o o Tel 1 440 337 1200 Tel 03 88 67 54 25 Tel 381 11 2622 945 Toll Free Tel 800 854 0530 Fax 03 88 67 19 45 Fax 381 11 2623 373 Fax 1 440 337 1180 Toll Free Fax 800 334 6999 Germany Singapore MP Biomedicals MP Biomedicals Singapore Europe Phone 0800 426 67337 Tel 65 6775 0008 Toll Free Phone Fax 0800 629 67337 Fax 65 6775 4536 00800 7777 9999 Toll Free Fax 00800 6666 8888 Switzerland Japan MP Biomedicals Switzerland Australia MP Bio Japan K K Tel 061 271 0007 MP Biomedicals Australasia Tel 03 3808 2102 Fax 061 271 0084 Pty Ltd Toll Free Tel 0120 788 020 Tel 61 2 9838 7422 Fax 03 3808 2401 United Kinad Fax 61 2 9838 7390 hited Raco MP Biomedicals UK Tel 0800 282 474 p The Netherlands Fax 0800 614 735 Belgium A MP Biomedicals Netherlands MP Biomedicals Tel 02 466 00 00 ira soo bone Fax 02 466 26 42 Canada Poland MP Biomedicals Canada MP Biomedicals Poland Tel 888 362 5487 Tel 48 22 659 58 95 Fax 514 935 7541 Fax 48 22 658 45 05 www mpbio com MP Biomedicals e 29525 Fountain Parkway e Solon OH 44139 e tel 1 800 854 0530 e fax 1 800 334 6999
7. nsiderations Before Use 3 1 Preparation of SEWS M Wash Solution 3 2 Precipitate Material in CLS TC Buffer 3 3 Sample Lysis with the FastPrep Instrument 3 4 Recovery of DNA from Dry Samples 3 9 Co Purification OT ANA orcos 4 Safety Precautions eeennne Bs FO csset pU SUEDE SIM DIUINIS 6 Example Data DNA Isolation from Animal and Plant Samples and Gel Electrophoresis 7 Table of Typical FastPrep Settings 8 Recommended Reference Format for Publication 9 HEIBIETICBS POCO e s c RUSO D CNN S NS andes 10 Related Products sseeeessesss 11 Product Use Limitation amp Warranty www mpbio com 1 Introduction to the FastDNA SPIN Green Kit and the FastPrep Instruments The FastDNA Green SPIN Kit quickly and efficiently isolates genomic DNA from a wide variety of animal and plant tissue sources Designed for use with the FastPrep Instruments from MP Biomedicals plant and animal tissues are easily lysed within 40 seconds These benchtop devices use a unique optimized motion to homogenize samples by multidirectional simultaneous impaction with lysing matrix particles FastPrep Instruments provide an extremely quick efficient and highly
8. rch or manufacturing use only They are not to be used as drugs or medical devices in order to diagnose cure mitigate treat or prevent diseases in humans or animals either as part of an accepted course of therapy or in experimental clinical investigation These products are not to be used as food food additives or general household items Purchase of MP Biomedicals products does not grant rights to reproduce modify or repackage the products or any derivative thereof to third parties MP Biomedicals makes no warranty of any kind expressed or implied including merchantability or fitness for any particular purpose except that the products sold will meet our specifications at the time of delivery Buyer s exclusive remedy and the sole liability of MP Biomedicals hereunder shall be limited to at our discretion no replacement or compensation product credits refund of the purchase price of or the replacement of materials that do not meet our specification By acceptance of the product Buyer indemnifies and holds MP Biomedicals harmless against and assumes all liability for the consequence of its use or misuse by the www mpbio com 13 FastDNA Green Spin Kit Buyer its employees or others including but not limited to the cost of handling Said refund or replacement is conditioned on Buyer notifying within thirty 30 days of receipt of product Failure of Buyer to give said notice within thirty 30 days shall constitute a waiver by
9. reproducible homogenization that surpasses traditional extraction methods using enzymatic digestion sonication blending douncing and vortexing Samples are placed into 2 0 ml tubes containing the proprietary Lysing Matrix D which are 1 4mm ceramic spheres While almost all samples are easily processed with Lysing Matrix D additional lysing matrix options are available at www mpbio com for more delicate samples or for tough hardened samples Homogenization in the FastPrep Instrument with Lysing Matrix D takes place in the presence of sample specific Cell Lysis Solutions CLS For plant tissues CLS VF is used in conjunction with a Protein Precipitation Solution PPS For all other samples including human and animal tissues CLS TC is used during sample lysis For maximum flexibility all buffers are provided in the kit Following lysis samples are centrifuged to pellet debris and lysing matrix DNA is purified from the supernatant with a silica based GENECLEAN procedure using SPIN filters Eluted DNA is ready for digestion electrophoresis PCR and any other desired application 2 Kit Components and User Supplied Materials 2 1 FastDNA Green SPIN Kit Components Lysing Matrix D 100x 2 0 ml tubes 1 4 Ceramic Spheres 100 spheres Binding Matrix 100 ml Concentrated SEWS M 12 ml DES 25 ml CLS VF 90 ml PPS 25 ml CLS TC 110 ml SPIN Modules 100 each Catch Tubes 100 each User manual 1 each MSDS 1 each Certificate of Analysis 1 e
10. sie som TT 00 oe Dua FE pem TT 6 EA ICO FI EC ET ETIN EA e CC mm TT E f oe oe ta m TT E m Du que m ETT E em Due pue pm TT eo sm EM 7 l PLANT oaa uere o oo E ae Laws e tang sowana 60 oe oras retenes soma usrowano ec oe oras matara Heuer amne sonoro es aose Caere p uere soma sowan er 9e Toese terrse omo somo 00 f ae Toese e AECI MET E roe Lum e 09 usen 69 ee Lure terrane tang sowan e ae Lomo e mre MEET E oe Lees tetas tong sowan e ae Duane ee tang ETT 00 em www mpbio com gt gt l l l FastPrep FastPrep EFE smono o0 ose IO e oo TT E mm Cimen ame omo TT e0 oe Lumens vona om TT 00 ETA Csen fe E TO TR CT NM Lum fe em CTO oo sm ICC fae o TT so oe ana fse o TO o0 oe ieoa fae Tm TT eo em Trina Sonam A om TO so A ICO ane omo TT e ane TEO ra eam TT 00 se menan termes remy TT 00 exse IE A omo TT 00 aan IEC E o TT E sm CO fee Cem TO E mem BACTERIA Teuna cw ET TO 00 ne RECTO oats woes TO o mee IEA cats ovens TO 60 ne sammas ots omues nsns 60 netus MEC oats wes TO p nones Teneo oats pomum O o mes Lumens oats mum TO ET YEAST AND FUNGI Estao rone ronse o cese ETT E 9 maanen cats reas somos 60 tse opines ca
11. st be secure but not over tightened to prevent sample leakage If the sample is too large for processing in a single tube divide the sample and process using multiple tubes MP Biomedicals Lysing Matrix particles and tubes have been rigorously tested and validated in the FastPrep Instrument The use of other products with the FastPrep Instrument is not recommended and may result in sample loss or instrument failure A single 40 second run at a speed setting of 6 0 in the FastPrep Instrument is sufficient to lyse almost all samples If the user experimentally determines that additional processing time is required the sample should be incubated on ice in the Lysing Matrix D tube for at least 2 minutes between successive FastPrep Instrument homogenizations to prevent overheating the sample and tube 6 www mpbio com MP Biomedicals recommends that all researchers begin the protocol with the Lysing Matrix D as supplied in the kit 1 4mm ceramic spheres If lysis is inefficient even after multiple runs of 40 seconds a single inch ceramic sphere provided can be added on top of the sample Depending on the sample lysis and or yield may or may not improve and shearing of existing genomic DNA may begin to occur Samples with the additional Ya inch ceramic sphere should be processed carefully in order to balance increased yield and lysis against increased DNA shearing by varying speed and or time settings 3 4 Recovery of DNA from Dry Sampl
12. t N 6035 050 50 preps e For use with yeast cells and fungal tissue e Lyse up to 10 cells per 2ml tube e Lysis and isolation with single phase organic solution in less than 90 minutes FastRNA Pro Green Kit Cat N 6045 050 50 preps e For use with all plant and animal samples e Lyse 50 100 mg tissue per 2ml tube e ysis and isolation with single phase organic solution in less than 90 minutes FastRNA Pro Soil Direct Kit and FastRNA Pro Soil Indirect Kit Cat N 6070 050 Cat N 6075 050 respectively 50 preps e solate RNA from soil samples direct kit and washed soil indirect kit in less than 2 hours e Variety of soil and environmental sample types e RNA protected during and after processing e Humic acids reduced to allow uninhibited RT PCR e ncludes additional reagents for even further purification if necessary e SPIN filters streamline silica handling FastProtein Blue Matrix Cat N 6550 400 50 preps Cat N 6550 500 100 preps e Release of proteins from gram positive and gram negative bacteria in 40 seconds e Protein extracts are ready for immediate electrophoresis or purification e deal for optimizing induction conditions FastProtein Red Matrix Cat N 6550 600 50 preps Cat N 6550 700 100 preps e Release of proteins from yeast cells and fungi in 40 seconds e Protein extracts are ready for immediate electrophoresis or purification e Ideal for optimizing induction conditions www mpbio co
13. the Buyer of all claims hereunder with respect to said material s FastDNA FastRNA FastPrep and BIO 1019 Systems are registered trademarks of MP Biomedicals LLC Take Advantage of FastPrep Kits Ready to use Protocols For DNA RNA And Protein Isolation From Any Sample e Rapid and reproducible sample lysis and purification process e No cross contamination with the closed lysing matrix tubes e Increased yields of high quality DNA RNA and proteins e Integrity and size of DNA RNA and proteins are retained e Nucleic acids and proteins are ready to use in downstream application DNA PROTEIN FastPrep Kits RNA FastDNA Kit and FastDNA Spin Kit Cat N 6540 400 Cat N 6540 600 respectively 100 preps e Lyse and isolate DNA in less than 30 minutes e Plant animal yeast fungal and microbial samples e No hazardous organic reagents required e SPIN filters streamline silica handling FastDNA Spin Kit FastDNA Spin Kit for Soil Cat N 6560 200 100 preps e yse and isolate DNA in less than 30 minutes e Variety of soil and environmental sample types e No hazardous organic reagents required e SPIN filters streamline silica handling 14 www mpbio com FastRNA Pro Blue Kit Cat N 6025 050 50 preps e For use with gram positive and gram negative bacteria e Lyse up to 10 cells per 2ml tube e Lysis and isolation with single phase organic solution in less than 90 minutes FastRNA Pro Red Kit Ca
14. ts ens somo 60 an ssa mani cats ens semanas 00 aras Chenmo cats reas TT 00 ETA www mpbio com 11 FastDNA Green Spin Kit 8 Recommended Reference Format for Publications DNA was isolated from specific sample using the FastDNA Green SPIN Kit and the FastPrep Instrument MP Biomedicals Santa Ana CA 9 References C albicans Sergey V Balashov et al 2006 Antimicrob Agents and Chemother Vol 50 2058 2063 Fungi P boydii Felix Gilgado et al 2005 J Clin Microbiol Vol 43 1930 1942 Fungi S schenckii Rita Marimon et al 2006 J Clin Microbiol Vol 44 3251 3256 Plant seeds Els J M Van Damme et al 2007 Plant Physiology Vol 144 662 672 Tomato leaves Hangsik Moon et al 2004 Journal of Experimental Botany Vol 55 402 1519 1528 Murine Intestine Alexandra J Scupham et al 2006 Appl Envir Microbiol Vol 72 793 801 Brain Jean E Jewell et al 2005 J Gen Virol Vol 86 2127 2134 Streptococcus pyogenes Audry C Almengor et al 2004 J Bacteriol Vol 186 7847 7857 Mycobacterium mucogenicum Gram positive bacteria Toidi Adekambi et al 2006 J Clin Microbiol Vol 44 837 840 Blood Jonas Bunikis et al 2004 J Infectious Disease Vol 189 1515 1523 12 www mpbio com 10 Related Products 11 Product Use Limitation Warranty The products presented in this instruction manual are for resea
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