FastDNA-96™ Fungal & Bacterial DNA Kit
Contents
1. 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 15 Pi Application Manual Revision 9696 400 11FEB Worldwide Ordering and Technical Support United States of America Worldwide Headquarters Tel 1 440 337 1200 Toll Free Tel 800 854 0530 Fax 1 440 337 1180 Toll Free Fax 800 334 6999 Europe Toll Free Phone 00800 7777 9999 Toll Free Fax 00800 6666 8888 Australia MP Biomedicals Australasia Pty Ltd Tel 61 2 9838 7422 Fax 61 2 9838 7390 Canada MP Biomedicals Canada Tel 888 362 5487 Fax 514 935 7541 France MP Biomedicals France Tel 03 88 67 54 25 Fax 03 88 67 19 45 Germany MP Biomedicals Gmbh Phone 0800 426 67337 Fax 0800 629 67337 Italy MP Biomedicals Italy Tel 0800 011643 Fax 0800 255 220 Japan MP Bio Japan K K Tel 03 3808 2102 Toll Free Tel 0120 788 020 Fax 03 3808 2401 The Netherlands MP Biomedicals Netherlands Tel 0800 0227416 _ Fax 0800 0227489 Poland MP Biomedicals Poland Tel 48 22 659 58 95 Fax 48 22 658 45 05 Russia MP Biomedicals Russia Tel 7 095 995 2844 Fax 7 095 995 2846 Serbia and Montenegro MP Global d o o Tel 381 11 2622 945 Fax 381 11 2623 373 Singapore a MP Biomedicals Asia Pacific2. of the Fungal Bacterial DNA Binding Solution 3 2 Binding Plate Pre Wash Buffer The FastDNA 96 Fungal amp Bacterial DNA Kit contains a bottle of Binding Plate Pre Wash Buffer which may form a precipitate over FastDNA 96 Fur time The precipitate is easily resuspended in solution by gently warming the bottle at 30 37 C for up to 30 minutes with mixing by inversion IMPORTANT Do not microwave this reagent 4 Safety Precautions Some of the supplied kit reagents contain 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 at all times when using this product Consult the Material Safety Data Sheet at www mpbio com for additional details This product is for research purposes only 5 Protocol NOTE Optimal reagent performance can be achieved by adding beta mercaptoethanol to the Fungal Bacterial DNA Binding Solution See Section 3 2 for details of this optimization 1 To the tubes of a FastDNA 96 Lysing Matrix Rack add 10 20 mg of wet weight bacterial or fungal cells resuspended in up to 50 ul of sterile water or sterile PBS Add 400ul of Lysis Buffer per well and re cap the tubes NOTE 10 20 mg of wet weight cultured cells or tissue is equivalent to the following 2x108 bacterial cells 2x107 yeast cells or 2x106 mammalian cells 2 Load the FastDNA 96 Lysing
3. MP 96 Binding Plate add 200ul of the Binding Plate Pre Wash Buffer Centrifuge the stacked plates for 5 minutes at 3 500 5 000 x g 10 To the wells of the MP 96 Binding Plate add 500 ul of Fungal Bacterial DNA Wash Buffer Centrifuge the stacked plates for 5 minutes at 3 500 5 000 x g 11 Stack the MP 96 Binding Plate atop a clean Elution Plate Add 50 100 ul of Elution Buffer 25ul minimum directly to the matrix inside the wells of the MP 96 Binding Plate Centrifuge the stacked plates for 5 minutes at 3 500 5 000 x g 12 Eluted DNA is now ready for PCR and other downstream applications To store the samples cover the Elution Plate with the supplied Foil Plate Cover Store samples at 4 C until use or at 20 C for extended periods 6 Example Data DNA Isolation from Bacteria amp Yeast Samples and Gel Electrophoresis MP Biomedicals Competitor Kit Yeast Yeast E coli Spores Spores E coli Figure 1 Comparison of DNA from S cerevisiae and E coli cells extracted with the FastDNA 96 Fungal amp Bacterial DNA Kit and a competitor kit Samples were loaded on a 0 8 agarose ethidium bromide gel 1 kb Ladder Marker NEB 7 Recommended Reference Format for Publications DNA was isolated from specific sample using the FastDNA 96 Fungal amp Bacterial DNA Kit and the FastPrep 96 Instrument MP Biomedicals LLC Santa Ana CA 8 Technical Support For technical support with this produ
4. Pte Ltd Tel 65 6775 0008 Fax 65 6775 4536 United Kingdom MP Biomedicals UK 1 Tel 0800 282 474 Ni Fax 0800 614 735 www mpbio com MP Biomedicals 29525 Fountain Parkway Solon OH 44139 e tel 1 800 854 0530 fax 1 800 334 6999
5. 7 y Application Manual Instruction Manual High Throughput Rapid Isolation of PCR Ready Genomic DNA from Tough to Lyse Fungi and Bacteria Samples using the FastPrep 96 System Catalog 9696 300 2 x 96 Preps Storage Ambient temperature 15 30 C Revision 9696 300 11FEB MP Biomedicals e 29525 Fountain Parkway e Solon OH 44139 e tel 1 800 854 0530 e fax 1 800 334 6999 FastDNA 96 Fungal amp Bacterial DNAS FastDNA 96 Fungal amp Bacterial DNA Kit High Throughput Rapid Isolation of PCR Ready Genomic DNA from Tough to Lyse Fungi and Bacteria Samples using the FastPrep 96 System Catalog 9696 300 2 x 96 Preps Storage Ambient temperature 15 30 C Revision 9696 300 11FEB FastDNA 96 Fungal amp TABLE OF CONTENTS 1 Introduction to the FastDNA 96 Fungal amp Bacterial DNA Kit and the FastPrep 96 InStrUMENt ooo ooo ooooooooooooooooo 5 2 Kit Components Storage amp User Supplied Materials 6 2 1 FastDNA 96 Fungal amp Bacterial DNA Kit Components 6 LD SUVA JEn mana omno eaaa a ngadang Aa gan E an GN ARR 7 2 3 User Supplied Materials o oo ocooooocoooooooooooooooo 7 3 Important Considerations Before Use o co o ccooooo cooooo coooo 7 3 1 Optimization of the Fungal Bacterial DNA Binding SOlUtION oo ooooocoooooooo oooooooooomomoma 7 3 2 Binding Plate Pre Wash Buffer ooo ocooooooooooooooooooooooo
6. MP Biomedicals harmless against and assumes all liability for the consequence of its use or misuse by the 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 the Buyer of all claims hereunder with respect to said material s GeneClean and FastPrep are registered trademarks of MP Biomedicals LLC 12 FastDNA 96 Fungal Other FastPrep Kits Available 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 Feces Cat N 6570 200 50 preps e Lyse and isolate PCR ready DNA in less than 30 minutes e Variety of stool sample types e No hazardous organic reagents required SPIN filters streamline silica handling FastDNA 50ml Spin Kit for Soil Cat N 6560 600 10 preps e Process low microbial containing soil samples e Lyse and isolate PCR ready DNA in less than 60 minutes e Variety of soil and environmental sample types e No hazardous organic reagents required e SPIN filters streamline silica handling FastRNA
7. Matrix Rack into the FastPrep 96 Instrument and process the samples A single 60 second run at a speed setting of 1600 rpm is sufficient to Iyse almost all samples If additional processing time is reguired over 5 minutes the FastDNA 96 Lysing Matrix Rack should be incubated on ice for at least 2 minutes between successive runs to prevent overheating the samples 3 Place the FastDNA 96 Lysing Matrix Rack in a microplate centrifuge adaptor and spin at 3 500 5 000 x g for 5 minutes 4 Transfer up to 250 ul of supernatant to the wells of a clean Deep Well Plate 5 Add 750 ul of Fungal Bacterial DNA Binding Solution to the supernatant in each well of the Deep Well Plate Cover the wells of the Deep Well Plate completely with the supplied Foil Plate Cover Place the samples on a plate shaker or vortexer and shake mix for 2 minutes 6 Centrifuge the Deep Well Plate for 5 minutes at 3 500 5 000 x g 7 Place an MP 96 Binding Plate on top of a supplied Collection Plate Remove the foil from the Deep Well Plate and transfer 500 ul of each supernatant to the wells of the MP 96 Binding Plate Centrifuge the stacked plates for 5 minutes at 3 500 5 000 x g 8 Discard the flow through from the Collection Plate and re use Repeat Step 7 until all of the supernatant has been carefully transferred to the binding plate 9 Continue to re use the Collection Plate by placing it beneath the MP 96 Binding Plate To the wells of the
8. Pro Blue Kit Cat N 6025 050 50 preps e For use with gram positive and gram negative bacteria e Lyse up to 1010 cells per 2ml tube e Lysis and isolation with single phase organic solution in less than 90 minutes 14 FastRNA Pro Red Kit Cat N 6035 050 50 preps e For use with yeast cells and fungal tissue e Lyse up to 1010 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 Lysis 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 Isolate RNA from soil samples direct kit and washed soil indirect kit in less than 2 hours e Variety of soil and environmental sample types RNA protected during and after processing e Humic acids reduced to allow uninhibited RT PCR e Includes additional reagents for even further purification if necessary 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 Ideal for optimizing induction conditions FastProtein
9. also be isolated if these hosts are present in your samples 2 Kit Components Storage and User Supplied Materials 2 1 FastDNA 96 Fungal amp Bacterial DNA Kit Components FastDNA 96 Lysing Matrix Rack 0 5 mm Beads 2 x 96 well rack Lysis Buffer 2x 40 ml Fungal Bacterial DNA Binding Solution 150 ml Binding Plate Pre Wash Buffer 50 ml Fungal Bacterial DNA Wash Buffer 100 ml Elution Solution 2x10 ml Deep Well Plate 2 each MP 96 Binding Plate 2 each Collection Plate 2 each Elution Plate 2 each Foil Plate Cover 4each User manual 1 each MSDS Online www mpbio com 1 each Certificate of Analysis 1 each 2 2 Storage All FastDNA 96 Fungal amp Bacterial DNA Kit reagents are stable at room temperature Storage should be maintained at room temperature The kit reagents are guaranteed for up to one year from the date of purchase of the kit 2 3 User Supplied Materials FastPrep 96 Instrument see Section 10 Centrifuge with a swing bucket style rotor that can spin up to 3 500 5 000 rpm Swing bucket centrifuge microplate adaptors for 96 well plates pair Plate shaker Sterile water beta mercaptoethanol 3 Important Considerations Before Use 3 1 Optimization of the Fungal Bacterial DNA Binding Solution Priorto beginning the Kit Protocolin Section 5 the Fungal Bacterial DNA Binding Solution can be optimized for increased reagent performance and yield Add 0 5 v v beta mercaptoethanol 750ul to the 150 ml bottle
10. ct please contact our MP Biomedical s Technical Support Team at 1 800 854 0530 by email at biotech mpbio com or visit us online at www mpbio com for live support For our European customers please contact our European Technical Support Team at 00 800 7777 9999 or by email at techsup eur mpbio com FastDNA 96 Fur 9 Product Use Limitation amp Warranty The products presented in this instruction manual are for research 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
11. o 7 4 Safety PpTECAUtIONS oo oooooooooooooooooooco WoWo Wo ooooo 8 Ds PEOLO GOL sasana anakan Ang ANG en amen an eg nangin ab aaa a kanan 8 6 Example Data DNA Isolation from Bacteria amp Yeast Samples and Gel ElectrophoreSis o ooooooooooooooooo ooooo ooooo 7 Recommended Reference Format for Publication 8 Technical SUpport w ooooooooooooocooo Woo ooo ooo ooomenaan 9 Product Use Limitation amp Warranty o oooooooooooooooocoooo 1 Introduction to the FastDNA 96 Fungal amp Bacterial DNA Kit and the FastPrep 96 Instrument The FastDNA 96 Fungal amp Bacterial DNA Kit quickly and efficiently isolates high quality PCR ready genomic DNA from tough to lyse fungi samples and bacteria in approximately 40 minutes The kit is designed for processing samples in 96 well plates and using the FastPrep 96 Instrument from MP Biomedicals Gram bacteria fungi including A fumigatus C albicans N crassa S cerevisiae S pombe amp mycelium as well as spores nematodes pollen and even mammalian cells are easily lysed in approximately 60 seconds The FastPrep 96 Instrument is a high throughput homogenizer developed to disrupt thoroughly any tissues and cells through the simultaneous bead beating and impaction of specialized Lysing Matrix beads on the sample material The FastPrep 96 Instrument uses a linear vertical bidirectional motion providing an extremely quick and highly reproducible homogenization that su
12. rpasses early generation homogenizers as well as traditional extraction methods using enzymatic digestion sonication blending douncing or vortexing Samples are placed into 1 2 ml size tubes of a FastDNA 96 Lysing Matrix Rack 96 deep well plate containing 0 5 mm lysing matrix beads The lysing matrix beads which are specially stabilized Zirconium oxide spheres are designed to efficiently lyse all organisms including historically difficult sources such as eubacterial spores and endospores gram bacteria fungi yeast molds and algae while in the presence of a specially formulated lysis solution The FastDNA 96 Fungal amp Bacterial DNA Kit isolates DNA from 10 20 mg of wet weight fungal or bacterial cells approximately 2x108 bacterial cells or 2x107 yeast cells or up to 40 mg soft animal tissue or Nematoda 2x106 mammalian cells Organic denaturants 5 FastDNA 96 Fungal amp or proteinases are not needed with this procedure Purified DNA is eluted in an EDTA free DNA elution solution and is ready for downstream applications including digestion electrophoresis and PCR A260 A280 ratios 1 8 Yield is typically 5 ug of total DNA eluted in 50 100 ul of elution solution 25ul minimum The FastDNA 96 Fungal amp Bacterial DNA Kit will recover genomic DNA fragments from 25 kb to 35 kb however some experiments have yielded results from as little as 100 bp up to gt 40 kb Viral and parasitic DNA will
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