taco DNARNA Extraction Kit User Manual RUO(201403)
Contents
1. Ensure that 135 ml 95 ethanol has been added to Washing Buffer A before the first time use Ensure that 230 ml 95 ethanol has been added to Washing Buffer B before the first time use 3 Magnetic Bead must be resuspended before aliquoting taco DNA RNA Extraction Kit Grind the tissue 40 mg with 450 ul PBS in a 1 5 ml micro centrifuge tube with disposable grinder Centrifuge at 12000 rpm for 5 minutes to spin down the debris For ethanol preserved sample please Appendix I Transfer 200 pl of the supernatant to column 1 7 of 96 Well Extraction Plate 48 Well Extraction Plate Open the door of taco to install the loaded plate and Mixing Sleeve Mixing Comb ensure they are pushed into position Note Please refer to respective instrument manual for instructions Close the door of taco and press Start Operation button to start extraction The extraction procedure will finished within hour After the extraction procedure take out the extraction plate and Mixing Sleeve Mixing Comb accordingly then press Reset Operation button to reset the program Note Please refer to respective instrument manual for instructions g Transfer the nucleic acids from column 6 and or 12 to the new micro centrifuge tubes for further use See Purity of 10 taco DNA RNA Extraction Kit Nucleic Acid Appendix II Note Carryover of magnetic beads in eluates will not affect most2. tace Research Use Only DNA RNA Extraction Kit User Manual Manufacturer GeneReach Biotechnology Corporation TEL 886 4 2463 9869 Email sales 9 genereach com No 19 Keyuan 2 Road Central Taiwan Science Park Taichung City 407 Taiwan Website www tacomag com 2014 03 taco DNA RNA Extraction Kit Content SVIMDOISsissssscscseadessescotencdoneceoncecssvebasessonsesesSvecdessdvosveceseosesessesseseosemesees 1 Kit COMPOMNeMt sis iscdcssccscscosicsstesseatescosscscdasssscessentessbecseatessonssvvdecssseese 2 As Reage Sonan et e tede ete teret oii ed Deme ina 2 B Gonsuniables 355 n iain tne si aia ens 3 MISI M 3 Materials and Equipments Required but Not Provided 4 Hore reo Oo PR ai 5 Intended Use 4 eee eee eee eee eee eene essen e et t etse ts stesse esses soo 5 Important NOt6S e 6 Product Limitations eee ee eee eee ee etes eene attestato aeta statuae 6 Nucleic Acid Extraction Procedure eee 8 A Use of taco Sticker for taco 32 ssssssssssss 8 B Protocol ett nette tee rere ele Yer er Eden 9 Troubleshoofig eee reor aerae Peor ieo ep rb noti oe reor o Pera avr eE Peer a oos 12 EVIL 16 Sample Preparation 0 eee ceeceseceseeeeeeeeeeeeaeeeseeceecaeseaeceaeenseees 16 taco DNA RNA Extraction Kit Appendix p
3. M 17 A Storage of Nucleic Acid seeeeeren 17 B Quantification of Nucleic Acid sess 17 C Purity of Nucleic Acid sar naiona tna a 17 taco DNA RNA Extraction Kit Symbols Date of manufacturing Manufacturer LOT Lot number Catalogue number Do not reuse E E G Kit Components A Reagents taco DNA RNA Extraction Kit taco DNA RNA Extraction Kit Cat No atc d rna Number of reactions 320 Reagent Name Volume Quantity Magnetic Bead Lysis Buffer Washing Buffer A Washing Buffer B Eluting Buffer User Manual 18 ml 1 bottle 180 ml 1 bottle 135 ml 2 bottles 40 ml 2 bottles 64 ml 1 bottle Note Treat all reagents as potential irritants Add 135 ml 95 ethanol to Washing Buffer A before use Mark the bottle label after adding ethanol Add 230 ml 95 ethanol to Washing Buffer B before use Mark the bottle label after adding ethanol taco DNA RNA Extraction Kit B Consumables Note Do not reuse the consumables a For taco 32 taco Plate amp Sleeve Product Name Amount pcs Cat No 96 Well Extraction Plate Mixing Sleeve taco Sticker b For taco 24 taco Plate amp Comb Product Name Amount pcs Cat No 48 Well Extraction Plate Mixing Comb 40 Storage All reagents should be sealed tightly in a cool and dry place at room temperature 16 30 C The expir
4. c Reagents were loaded in wrong order d Poor sample quality e Incorrect sample volume f Mixing Sleeve Mixing Comb was not installed g Inappropriate operation environment taco DNA RNA Extraction Kit Comments and suggestions Restart the loading procedure with a new extraction plate Ensure that all reagents were loaded on the well in the correct order Repeat the extraction procedure with new samples Using fresh sample for extraction is recommended Poor sample quality may influence nucleic acid quality The kit performance would be affected if user did not use the right volume of sample User should optimize the sample quantity when dealing with different sample types Contact your local distributor or GeneReach Biotechnology Corporation for assistance Operation temperature could affect the recovery rate Please ensure the operation environment is under room temperature 16 30 C 13 h Use non recommended extraction instrument taco DNA RNA Extraction Kit Comments and suggestions Using non recommended instrument may influence the performance of taco DNA RNA Extraction Kit We strongly recommend user to apply DNA RNA Extraction Kit on taco extraction systems Poor DNA RNA performance in downstream applications Note A spectrophotometer is required for following check up a Insufficient DNA RNA is used in downstream application b Excess DNA RNA u
5. 2014 GeneReach Biotechnology Corporation All rights reserved For research use only Not intended for any animal or human therapeutic or diagnostic use 18
6. downstream applications f the risk of magnetic beads carryover needs to be minimized transfer the eluates to micro centrifuge tubes centrifuge for 1 minute at full speed to pellet down the remaining magnetic beads and carefully transfer the supernatants to new micro centrifuge tubes h It is strongly recommended to use freshly extracted nucleic acids for downstream applications such as amplification Otherwise the extracted nucleic acids should be kept frozen for long term storage 80 C is recommended See Storage of Nucleic Acid Appendix II Note Do not reuse the Consumables Note Any deviation from the instruction may lead to a low recovery rate of the nucleic acid extract Troubleshooting Low DNA RNA yield a Magnetic bead was not resuspended completely b Washing Buffer A and B did not contain ethanol taco DNA RNA Extraction Kit Comments and suggestions Before starting the procedure ensure that magnetic bead is fully resuspended Vortex for at least 5 seconds before first use and perform mild agitation before subsequent uses Ensure the correct volume of ethanol is added to Washing Buffer A and B tightly seal the reagent bottles to prevent ethanol from evaporating Repeat the extraction procedure with proper reagent is necessary when the ethanol was not added to Wash Buffer A and Wash Buffer B before use For the proper procedure of extraction please see Protocol 12
7. ation date of the kit and each component are stated on the label of each item Do not use any reagent of the kit beyond the expiration date Users should check the expiration date before use as it could affect the accuracy of the result taco DNA RNA Extraction Kit Materials and Equipments Required but Not Provided taco Nucleic Acid Automatic Extraction System taco 24 taco 32 or taco mini Step pipette optional Disposable gloves Micro centrifuge tubes Micropipette and filter tips p1000 p200 e 95 ethanol Phosphate buffer saline PBS taco DNA RNA Extraction Kit Introduction The taco DNA RNA Extraction Kit is designed for taco Nucleic Acid Automatic Extraction System Based on the magnetic separation technology homogenized sample cells are lysed and nucleic acid is captured by silica coated magnetic beads Washing Buffer is then applied to remove impurities and Eluting Buffer to recover nucleic acids from magnetic beads following serial washing steps This kit can extract viral DNA and RNA from shrimp muscle for research use purpose only Other sample types must be validated by users Intended Use The taco DNA RNA Extraction Kit is intended to be used for extracting viral DNA and RNA from various sample types such as shrimp tissue The taco DNA RNA Extraction Kit has to be used with the taco Nucleic Acid Automatic Extraction System This product is inten
8. ded to be used by professional users such as well trained laboratory technicians who are familiar with molecular biology techniques taco DNA RNA Extraction Kit Important Notes After receiving the kit please check the kit components for any damage Contact GeneReach Biotechnology Corporation or your local distributor if reagent bottles are damaged Do not use damaged kit as it could affect the accuracy of the result Pipette tips are all for one time use only Repeated usage will lead to cross contamination When working with chemicals please always wear a suitable lab coat disposable gloves and protective goggles Discard gloves if they are contaminated Do not combine components with different batches Avoid microbial contamination of the reagents To minimize the risk of infections from potentially infectious material we recommend working under a laminar hood until the samples are lysed This kit should only be used by trained personnel Disposal of waste must be compliant with local laws Product Limitations The system performance has been validated by using infected shrimp muscle for viral nucleic acid isolation The user is responsible for taco DNA RNA Extraction Kit validating the performance of the taco DNA RNA Extraction Kit for other particular use The kit and plastic parts are not intended for any therapeutic or diagnostic purposes for animals or humans taco DNA RNA Extraction Kit Nuc
9. leic Acid Extraction Procedure A Use of taco Sticker for taco 32 For your convenience you may put the taco Sticker on top of reagent bottles and on the rim of 96 Well Extraction Plate to avoid human error a taco M Sticker Plate Sticker For taco 32 Apply the Plate Sticker on the rim of 96 Well Extraction Plate directly LB wam wa BRE c ie we NENNEN E Bottle Sticker Apply the Sticker on top of each reagent bottle ZR di ida LB M WA WA moms QE b Abbreviation Definition LB Lysis Buffer M Magnetic Bead WA Washing Buffer A WAM Washing Buffer A Magnetic Bead WB Washing Buffer B E Eluting Buffer taco DNA RNA Extraction Kit B Protocol Note The following protocol is for fresh and frozen shrimp samples For other sample types please refer to www tacomag com for instructions a Load reagents into 96 Well Extraction Plate 48 Well Extraction Plate according to Table 1 at the room temperature 16 30 C for the best performance Table 1 Loading Reagent Step Reagents Add 200 pl 95 ethanol and 500 pl Lysis Buffer to column 1 7 2 Add 750 pl Washing Buffer A to column 2 8 3 Add 750 pl Washing Buffer A to column 3 9 4 Add 750 ul Washing Buffer B to column 4 10 5 Add 750 ul Washing Buffer B to column 5 11 6 Add 50 200 pl Eluting Buffer to column 6 12 7 Add 50 pl Magnetic Bead to column 2 8
10. se refer to www tacomag com 16 taco DNA RNA Extraction Kit Appendix II A Storage of Nucleic Acid For long term storage extracted nucleic acids should be stored at 80 C B Quantification of Nucleic Acid Note A spectrophotometer is required for following check up The concentration of nucleic acids should be determined by measuring the absorbance at 260 nm in a spectrophotometer Use Hluting Buffer as the blank to calibrate the spectrophotometer If the purified nucleic acids need to be diluted before the quantification the Eluting Buffer also has to be diluted before use Also the same dilution factor needs to be applied for calculation Collect the absorbance reading of purified nucleic acid at 260 nm and 280 nm The ratio between the absorbance values at 260 nm and 280 nm gives an estimation of nucleic acid purity See Purity of Nucleic Acid Carryover of magnetic beads may affect the A26 reading but should not affect the performance of nucleic acid in downstream applications C Purity of Nucleic Acid Purity is determined by calculating the ratio of absorbance at 260 nm to absorbance at 280 nm with a background correction at 320 nm i e A260 A320 A280 A320 A subtracted absorbance reading at 320 17 taco DNA RNA Extraction Kit nm is to correct the presence of magnetic beads particles in the eluted solution An A260 A so ratio of 1 6 2 0 is indicative of highly purified nucleic acid
11. sed in downstream application Quantify the extracted DNA RNA by spectrophotometer of the absorbance at 260 nm See Quantification of Nucleic Acid Appendix II Excess DNA RNA can inhibit some enzymatic reactions Quantify the extracted DNA RNA by spectrophotometer of the absorbance at 260 nm See Quantification of Nucleic Acid Appendix II 14 Low A260 A 280 ratio a Absorbance reading at 320 nm was not subtracted from the absorbance readings at 260 nm and 280 nm taco DNA RNA Extraction Kit Comments and suggestions To correct for the presence of magnetic beads particles in the eluted solution an absorbance reading at 320 nm should be taken and subtracted from the absorbance readings obtained at 260 nm and 280 nm 15 taco DNA RNA Extraction Kit Appendix I Sample Preparation B Animal Tissue ethanol preserved shrimp muscle i lil iv Grind the tissue 20 mg with 500 ul Lysis Buffer in 1 5 ml micro centrifuge tube with disposable grinder Centrifuge at 12000 rpm for 5 minutes to spin down the debris Transfer 400 ul of the supernatant and 200 ul 95 ethanol to column 1 7 of 96 Well Extraction Plate Follow Table 1 from Step 2 to 7 for loading reagents Note The above sample preparation method is recommended for general muscle tissues which contain high volume of protein other sample types need to be validated by users Note For other sample types plea
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