NucleoSpin® Plasmid EasyPure - MACHEREY
Contents
1. Plasmid DNA purification User manual NucleoSpin Plasmid EasyPure July 2014 Rev 02 MACHEREY NAGEL www mn net com Plasmid DNA purification Protocol at a glance Rev 02 NucleoSpin Plasmid EasyPure 1 Cultivate and harvest bacterial 12 000 x g cells c 30s 2 Cell lysis 150 uL Buffer A1 250 uL Buffer A2 RT up to 2 min 350 uL Buffer A3 3 Clarification of the lysate e 212 000 x g 3 min Bind DNA a er Load supernatant S ce 1 000 2 000 x g D 30s 5 Wash and dry silica F membrane 450 uL Buffer AQ S 5 gt 12 000 x g ej 1 min 6 Elute DNA 50 uL Buffer AE e RT 1 min gt 12 000 x g 1 min MACHEREY NAGEL GmbH amp Co KG Neumann Neander Str 6 8 52355 D ren Germany Tel 49 24 21 969 270 Fax 49 24 21 969 199 tech bio mn net com www mn net com Plasmid DNA purification Table of contents 1 Components 4 1 1 Kit contents 4 1 2 Reagents consumables and equipment to be supplied by user 5 1 3 About this user manual 5 2 Product description 6 2 1 Basic principle 6 2 2 Kit specifications 6 2 3 Growth of bacterial cultures 7 2 4 Elution procedures 8 3 Storage conditions and preparation of working solutions 10 4 Safety instructions 11 5 NucleoSpin Plasmid EasyPure protocol 13 6 Appendix 15 6 1 Troubleshooting 15 6 2 Ordering information 18 6 3 References 18 6 4 Product use restriction warranty 19 MACHEREY NAGEL 07 2012. 07 2010 Rev 03 Please contact MACHEREY NAGEL GmbH amp Co KG Tel 49 24 21 969 270 tech bio 2 mn net com 20 MACHEREY NAGEL 07 2014 Rev 02
3. revenue or profit whether based upon warranty contract tort including negligence or strict liability arising in connection with the sale or the failure of MACHEREY NAGEL products to perform in accordance with the stated specifications This warranty is exclusive and MACHEREY NAGEL makes no other warranty expressed or implied The warranty provided herein and the data specifications and descriptions of this MACHEREY NAGEL product appearing in MACHEREY NAGEL published catalogues and product literature are MACHEREY NAGEL s sole representations concerning the product and warranty No other statements or representations written or oral by MACHEREY NAGEL s employees agent or representatives except written statements signed by a duly authorized officer of MACHEREY NAGEL are authorized they should not be relied upon by the customer and are not a part of the contract of sale or of this warranty Product claims are subject to change Therefore please contact our Technical Service Team for the most up to date information on MACHEREY NAGEL products You may also contact your local distributor for general scientific information Applications mentioned in MACHEREY NAGEL literature are provided for informational purposes only MACHEREY NAGEL does not warrant that all applications have been tested in MACHEREY NAGEL laboratories using MACHEREY NAGEL products MACHEREY NAGEL does not warrant the correctness of any of those applications Last updated
4. NAGEL 07 2014 Rev 02 11 Plasmid DNA purification Precaution phrases P 3014312 P 3024352 P 304 340 P 305 351 338 P 330 P 3324313 P 333 313 P 337 313 P 3424311 P 363 P 390 P 406 IF SWALLOWED Call a POISON CENTER doctor if you feel unwell BEI VERSCHLUCKEN Bei Unwohlsein GIFTINFORMATIONSZENTRUM Arzt anrufen IF ON SKIN Wash with plenty of water BEI KONTAKT MIT DER HAUT Mit viel Wasser waschen IF INHALED Remove victim to fresh air and keep at rest in a position com fortable for breathing BEI EINATMEN An die frische Luft bringen und in einer Position ruhigstellen die das Atmen erleichtert IF IN EYES Rinse continuously with water for several minutes Remove contact lenses if present and easy to do continue rinsing BEI KONTAKT MIT DEN AUGEN Einige Minuten lang behutsam mit Wasser sp len Vorhandene Kontaktlinsen nach M glichkeit entfernen Weiter sp len Rinse mouth Mund aussp len If skin irritation occurs Get medical advice attention Bei Hautreizung Arztlichen Rat einholen rztliche Hilfe hinzuziehen If skin irritation or a rash occurs Get medical advice attention Bei Hautreizung oder ausschlag Arztlichen Rat einholen rztliche Hilfe hinzuziehen Get medical advice attention Bei anhaltender Hautreizung Arztlichen Rat einholen rztliche Hilfe hinzuziehen If experiencing respiratory symptoms Call a POISON CENTER doctor Bei Sy
5. Usually an OD of 3 6 can be achieved Alternatively rich media like 2x YT Yeast Tryptone TB Terrific Broth or CircleGrow can be used In this case bacteria grow faster reach the stationary phase much earlier than in LB medium x12 h and higher cell masses can be reached However this does not necessarily yield more plasmid DNA Overgrowing a culture might lead to a higher percentage of dead or starving cells and the resulting plasmid DNA might be partially degraded or contaminated with chromosomal DNA To find the optimal culture conditions the culture medium and incubation times have to be optimized for each host strain plasmid construct combination individually Cell cultures should be grown under antibiotic selection at all times to ensure plasmid propagation Without this selective pressure cells tend to lose a plasmid during cell division Since bacteria grow much faster without the burden of a high copy plasmid they take over the culture rapidly and the plasmid yield goes down regardless of the cell mass Table 2 gives information on concentrations of commonly used antibiotics Hands on time MACHEREY NAGEL 07 2014 Rev 02 7 Plasmid DNA purification Table 2 Information about antibiotics according to Maniatis Antibiotic Stock solution Storage Working concentration concentration Ampicillin 50 mg mL in H O 20 C 20 50 ug mL Carbenicillin 50 mg mL in H O 20 C 20 60 ug mL Chloramphenicol 34 mg mL in Et
6. 4 Rev 02 3 Plasmid DNA purification 1 Components 1 1 Kit contents NucleoSpin Plasmid EasyPure 10 preps 50 preps 250 preps REF 740727 10 740727 50 740727 250 Resuspension Buffer A1 5mL 15 mL 75 mL Lysis Buffer A2 5mL 15 mL 100 mL Neutralization Buffer A3 5mL 20 mL 100 mL Wash Buffer AM 6 mL 6mL 25 mL Concentrate Elution Buffer AE 13 mL 13 mL 30 mL Liquid RNase A 2mg 6 mg 30 mg NucleoSpin Plasmid EasyPure Columns 10 50 250 dark blue rings Collection Tubes 2 mL 10 50 250 Short protocol 1 1 1 For preparation of working solutions and storage conditions see section 3 Composition of Elution Buffer AE 5 mM Tris HCl pH 8 5 4 MACHEREY NAGEL 07 2014 Rev 02 Plasmid DNA purification 1 2 Reagents consumables and equipment to be supplied by user Reagents 96 100 ethanol Consumables 1 5 mL microcentrifuge tubes for sample lysis and DNA elution Disposable pipette tips Equipment Manual pipettors Centrifuge for microcentrifuge tubes e Vortex mixer Personal protection equipment lab coat gloves goggles 1 3 About this user manual It is strongly recommended reading the detailed protocol sections of this user manual if the NucleoSpin Plasmid EasyPure kit is used for the first time Experienced users however may refer to the Protocol at a glance instead The Protocol at a glance is designed to be used only as a supplemental tool for quick referencing while performing the purif
7. C to dry silica membrane completely Elute DNA Place the NucleoSpin Plasmid EasyPure Column in a 1 5 mL microcentrifuge tube not provided and add 50 uL Buffer AE Incubate for 1 min at room temperature 18 25 C Centrifuge for 1 min at full speed 7 12 000 x g Note For more efficient elution procedures and alternative elution buffer e g TE buffer or water see section 2 4 Load supernatant 1 000 2 000 x g 30 s 450 pL AQ gt 12 000 x g 1 min 50 uL AE RT 1 min gt 12 000 x g 1 min MACHEREY NAGEL 07 2014 Rev 02 Plasmid DNA purification 6 Appendix 6 1 Troubleshooting Problem Possible cause and suggestions Cell pellet not properly resuspended It is essential that the cell pellet is completely resuspended prior to lysis No cell clumps should be visible before addition of Buffer A2 dei SDS in Buffer A2 precipitated in Storage of Buffer A2 below 20 C may cause precipitation of cells SDS If salt precipitate is observed incubate buffer at 30 40 C for several minutes and mix well until all precipitate is redissolved completely Cool down to room temperature before use Too many bacterial cells used See table 3 for maximum amount of cells Incomplete lysis of bacterial cells See Possible cause and suggestions above No or insufficient amounts of antibiotic used during cultivation Cells carrying the plasmid of interest may become overgrown by
8. OH 20 C 25 170 ug mL Kanamycin 10 mg mL in H O 20 C 10 50 ug mL Streptomycin 10 mg mL in H O 20 C 10 50 ug mL Tetracycline 5 mg mL in EtOH 20 C 10 50 g mL As rule of thumb use 5 mL of a well grown LB culture as given in the kit specifications However the culture volume can be increased if the cell culture grows very poorly or has to be decreased if e g very rich culture media were used Refer to Table 3 to choose the best culture volume according to the optical density at 600 nm OD Table 3 Recommended culture volumes for NucleoSpin Plasmid EasyPure Culture volume Note if too much bacterial material is used the lysis and precipitation steps become inefficient causing decreased yield and plasmid quality If more than the recommended amount of cells shall be processed increase all lysis buffers proportionally 2 4 Elution procedures The elution buffer volume and method can be adapted to the subsequent downstream application to achieve higher yield and or concentration than the standard method recovery about 70 90 Higher yield in general especially for larger constructs Heat elution buffer to 70 C add 50 100 uL to the NucleoSpin Plasmid EasyPure Column and incubate at 70 C for 2 min High yield Perform two elution steps with the volume indicated in the individual protocol About 90 100 of bound nucleic acids can be eluted Maniatis T Fritsch EF Sambrook J Molecular clonin
9. Y FORBIDDEN The respective user is liable for any and all damages resulting from such application DNA RNA PROTEIN purification products of MACHEREY NAGEL are suitable for IN VITRO USES ONLY ONLY MACHEREY NAGEL products specially labeled as IVD are also suitable for IN VITRO diagnostic use Please pay attention to the package of the product IN VITRO diagnostic products are expressly marked as IVD on the packaging IF THERE IS NO IVD SIGN THE PRODUCT SHALL NOT BE SUITABLE FOR IN VITRO DIAGNOSTIC USE ALL OTHER PRODUCTS NOT LABELED AS IVD ARE NOT SUITED FOR ANY CLINICAL USE INCLUDING BUT NOT LIMITED TO DIAGNOSTIC THERAPEUTIC AND OR PROGNOSTIC USE No claim or representations is intended for its use to identify any specific organism or for clinical use included but not limited to diagnostic prognostic therapeutic or blood banking It is rather in the responsibility of the user or in any case of resale of the products in the responsibility of the reseller to inspect and assure the use of the DNA RNA protein purification products of MACHEREY NAGEL for a well defined and specific application MACHEREY NAGEL shall only be responsible for the product specifications and the performance range of MN products according to the specifications of in house quality control product documentation and marketing material This MACHEREY NAGEL product is shipped with documentation stating specifications and other technical information MACHERE
10. Y NAGEL warrants to meet the stated specifications MACHEREY NAGEL s sole obligation and the customer s sole remedy is limited to replacement of products free of charge in the event products fail to perform as warranted Supplementary reference is made to the general business terms and conditions of MACHEREY NAGEL which are printed on the price list Please contact us if you wish to get an extra copy There is no warranty for and MACHEREY NAGEL is not liable for damages or defects arising in shipping and handling transport insurance for customers excluded or out of accident or improper or abnormal use of this product defects in products or MACHEREY NAGEL 07 2014 Rev 02 19 Plasmid DNA purification components not manufactured by MACHEREY NAGEL or damages resulting from such non MACHEREY NAGEL components or products MACHEREY NAGEL makes no other warranty of any kind whatsoever and SPECIFICALLY DISCLAIMS AND EXCLUDES ALL OTHER WARRANTIES OF ANY KIND OR NATURE WHATSOEVER DIRECTLY OR INDIRECTLY EXPRESS OR IMPLIED INCLUDING WITHOUT LIMITATION AS TO THE SUITABILITY REPRODUCTIVITY DURABILITY FITNESS FOR A PARTICULAR PURPOSE OR USE MERCHANTABILITY CONDITION OR ANY OTHER MATTER WITH RESPECT TO MACHEREY NAGEL PRODUCTS In no event shall MACHEREY NAGEL be liable for claims for any other damages whether direct indirect incidental compensatory foreseeable consequential or special including but not limited to loss of use
11. cells without plasmid see table 2 when inadequate levels of the antibiotics are used Add appropriate amounts of freshly Poor prepared stock solutions to all media both solid and liquid plasmid yield Bacterial culture too old Do not incubate cultures for more than 16h LB or 12h rich media at 37 C under shaking to avoid starvation and plasmid degradation Incomplete neutralization Mix thoroughly after addition of Buffer A3 until LyseControl has turned colorless without any traces of blue MACHEREY NAGEL 07 2014 Rev 02 15 Plasmid DNA purification Suboptimal elution conditions f possible use a slightly alkaline elution buffer like Buffer AE 5 M Tris HCl pH 8 5 If nuclease free water is used check the Poor pH of the water Elution efficiencies drop drastically with buffers plasmid lt pH 7 yield continued Low copy number plasmid was used Atleast double or triple culture volume and increase lysis buffers if final amount of cells exceed the recommended volumes of table 3 Reagents not applied properly Add indicated volume of 96 100 ethanol to Buffer AQ Concentrate and mix thoroughly see section 3 Nuclease rich host strains used f Especially when working with nuclease rich strains keep No plasmid plasmid preparations on ice or frozen in order to avoid DNA yield degradation Inappropriate storage of plasmid DNA Quantitate DNA directly after preparation for example by agarose g
12. el electrophoresis Store plasmid DNA dissolved in water at lt 18 C or at lt 5 C when dissolved in Buffer AE or TE buffer Nicked plasmid DNA Cell suspension was incubated with alkaline Lysis Buffer A2 too long Reduce lysis time Poor Genomic DNA contamination plasmid Cell lysate was vortexed or mixed too vigorously after addition quality of Buffer A2 Genomic DNA was sheared and thus liberated Smeared plasmid bands on agarose gel Especially when working with nuclease rich strains keep plasmid preparations on ice or frozen in order to avoid DNA degradation 16 MACHEREY NAGEL 07 2014 Rev 02 Plasmid DNA purification Suboptimal performance of plasmid DNA in enzymatic reactions Carry over of ethanol Make sure that the NucleoSpin Plasmid EasyPure Column is completely dry after step 5 Otherwise discard flow through and repeat centrifugation Elution of plasmid DNA with TE buffer EDTA may inhibit sequencing reactions Repurify plasmid DNA and elute with Buffer AE or water Alternatively the eluted plasmid DNA can be precipitated with ethanol and redissolved in Buffer AE or water Not enough DNA used for sequencing reaction Quantify DNA by agarose gel electrophoresis before setting up sequencing reactions MACHEREY NAGEL 07 2014 Rev 02 17 Plasmid DNA purification 6 2 Ordering information Product REF NucleoSpin Plasmid EasyPure 740727 10 50 250 foie ate plas
13. eoSpin Plasmid EasyPure Column features a new specially treated silica membrane which allows speeding up the procedure by a combined washing and drying step No additional steps are necessary if nuclease rich host strains are used The number of washing and drying steps is reduced from 3 to only 1 LyseControl The Lysis Buffer A2 contains a blue pH indicator to ensure complete neutralization for maximum yield The purified plasmid DNA is suitable for applications like automated fluorescent DNA sequencing PCR or any kind of enzymatic manipulation For preparation of working solutions and storage conditions see section 3 Composition of Elution Buffer AE 5 mM Tris HCl pH 8 5 6 MACHEREY NAGEL 07 2014 Rev 02 Plasmid DNA purification Table 1 Kit specifications at a glance Parameter NucleoSpin Plasmid EasyPure Culture volume 2 10 mL Typical yield 15 30 ug Elution volume 50 uL Binding capacity 35 ug Vectors 15 kbp Preparation time 14 min 6 preps Format Mini spin column 2 3 Growth of bacterial cultures Yield and quality of plasmid DNA highly depend on the type of culture media and antibiotics the bacterial host strain the plasmid type size or copy number For cultivation of bacterial cells harbouring standard high copy plasmids we recommend LB Luria Bertani medium The cell culture should be incubated at 37 C with constant shaking 200 250 rpm preferably 12 16 h over night
14. g A laboratory manual Cold Spring Harbor Cold Spring New York 1982 8 MACHEREY NAGEL 07 2014 Rev 02 Plasmid DNA purification High concentration Perform one elution step with 60 of the volume indicated in the individual protocol Concentration of DNA will be higher than with standard elution approx 130 96 Maximal yield of bound nucleic acids is about 80 High yield and high concentration Apply half of the volume of elution buffer as indicated in the individual protocol incubate for 3 min and centrifuge Apply a second aliquot of elution buffer incubate and centrifuge again Thus about 85 100 of bound nucleic acids are eluted with the standard elution volume at a high concentration Elution Buffer AE 5 mM Tris HCl pH 8 5 can be replaced by TE buffer or water as well However we recommend using a weakly buffered slightly alkaline buffer containing no EDTA especially if the plasmid DNA is intended for sequencing reactions If water is used the pH should be checked and adjusted to pH 8 0 8 5 since deionized water usually exhibits a pH below 7 Furthermore absorption of CO leads to a decrease in pH of unbuffered solutions MACHEREY NAGEL 07 2014 Rev 02 9 Plasmid DNA purification 3 Storage conditions and preparation of working solutions Attention Buffer A3 contains guanidine hydrochloride Wear gloves and goggles CAUTION Buffer A3 contains guanidine hydrochloride which can form highly reac
15. ication procedure All technical literature is available on the internet at www mn net com Please visit the MACHEREY NAGEL website to verify that you are using the latest revision of this user manual Please contact Technical Service regarding information about changes of the current user manual compared to previous revisions MACHEREY NAGEL 07 2014 Rev 02 5 Plasmid DNA purification 2 Product description 2 1 Basic principle With the NucleoSpin Plasmid EasyPure method the pelleted bacteria are resuspended Buffer A1 and plasmid DNA is liberated from the E coli host cells by SDS alkaline lysis Buffer A2 Buffer A3 neutralizes the resulting lysate and creates appropriate conditions for binding of plasmid DNA to the silica membrane of the NucleoSpin Plasmid EasyPure Column Precipitated protein genomic DNA and cell debris are then pelleted by a centrifugation step The supernatant is loaded onto a NucleoSpin Plasmid EasyPure Column With the NucleoSpin Plasmid EasyPure kit contaminations like salts metabolites nucleases and soluble macromolecular cellular components are removed by only a single washing step with Buffer AQ Pure plasmid DNA is finally eluted under low ionic strength conditions with slightly alkaline Buffer AE 5 mM Tris HCl pH 8 5 2 2 Kit specifications The NucleoSpin Plasmid EasyPure kits are designed for the rapid small scale preparation of highly pure plasmid DNA mini preps The Nucl
16. l precipitate is dissolved completely Cool buffer down to room temperature 18 25 C before use y Mix Add 250 uL Buffer A2 Mix gently by inverting the tube RT 2 min 5 times Do not vortex to avoid shearing of genomic DNA Incubate at room temperature 18 25 C for up to 2 min or until lysate appears clear Add 350 uL Buffer A3 Mix thoroughly by inverting the E SD0 UE AS tube until LyseControl has turned colorless throughout Mix the lysate without any traces of blue color Do not vortex to avoid shearing of genomic DNA Clarification of lysate y Centrifuge for 3 min at full speed gt 12 000 x g gt 12 000 x g e 3 min MACHEREY NAGEL 07 2014 Rev 02 13 NucleoSpin Plasmid EasyPure Bind DNA Place a NucleoSpin Plasmid EasyPure Column into a Collection Tube 2 mL and decant the supernatant from step 3 onto the column Centrifuge for 30 s at 1 000 2 000 x g Discard flow through and place the spin column back into the collection tube Wash and dry silica membrane Add 450 uL Buffer AQ supplemented with ethanol see section 3 Centrifuge for 1 min at full speed gt 12 000 x g Very carefully discard the collection tube and the flow through and make sure the spin cup outlet does not touch the wash buffer surface Otherwise repeat the centrifugation step Note To reduce ethanol carry over to a minimum for better performance in downstream applications incubate spin cup for 10 15 min at 37
17. mids 740958 Buffer A2 740912 1 Buffer A3 740913 1 EUM ca ka a 740995 Buffer AE 740917 1 Liquid RNase A 740397 Collection Tubes 2 mL 740600 6 3 References Pack of 10 50 250 preps 1 1L 1L 1L 20 mL 1L 250 mg 1000 Birnboim H C and J Doly 1979 A rapid alkaline extraction procedure for screening of recombinant plasmid DNA Nucleic Acids Res 7 1513 1523 Vogelstein B and D Gillespie 1979 Preparative and analytical purification of DNA from agarose Proc Natl Acad Sci USA 76 615 619 18 MACHEREY NAGEL 07 2014 Rev 02 Plasmid DNA purification 6 4 Product use restriction warranty NucleoSpin Plasmid EasyPure kit components are intended developed designed and sold FOR RESEARCH PURPOSES ONLY except however any other function of the product being expressly described in original MACHEREY NAGEL product leaflets MACHEREY NAGEL products are intended for GENERAL LABORATORY USE ONLY MACHEREY NAGEL products are suited for QUALIFIED PERSONNEL ONLY MACHEREY NAGEL products shall in any event only be used wearing adequate PROTECTIVE CLOTHING For detailed information please refer to the respective Material Safety Data Sheet of the product MACHEREY NAGEL products shall exclusively be used in an ADEQUATE TEST ENVIRONMENT MACHEREY NAGEL does not assume any responsibility for damages due to improper application of our products in other fields of application Application on the human body is STRICTL
18. mptomen der Atemwege GIFTINFORMATIONSZENTRUM Arzt anrufen Wash contaminated clothing before reuse Kontaminierte Kleidung vor erneutem Tragen waschen Absorb spillage to prevent material damage Versch ttete Menge aufnehmen um Materialsch den zu vermeiden Store in a corrosive resistant container with a resistant inner liner In korrosionsbest ndigem Beh lter mit korrosionsbest ndiger Auskleidung aufbewahren For further information please see Material Safety Data Sheets www mn net com Weiterf hrende Informationen finden Sie in den Sicherheitsdatenbl ttern www mn net com 12 MACHEREY NAGEL 07 2014 Rev 02 NucleoSpin Plasmid EasyPure 5 NucleoSpin Plasmid EasyPure protocol Before starting the preparation Check if Wash Buffer AQ was prepared according to section 3 Cultivate and harvest bacterial cells Use 2 10mL of a saturated E coli culture see page 8 table 3 pellet cells in a standard benchtop microcentrifuge for 30 s at 12 000 x g e 212 000 x g Discard the supernatant and remove as much of the 30s liquid as possible Cell lysis 150 pL A1 Add 150yuL Buffer A1 Resuspend the cell pellet ii completely by vortexing or pipetting up and down Make Resuspend sure no cell clumps remain before addition of Buffer A2 Attention Check Buffer A2 for precipitated SDS If a white precipitate is visible warm the buffer for several minutes 250 uL A2 at 30 40 C unti
19. rases up to 125 mL or 125 g Mindergef hrliche Eigenschaften m ssen bis 125 mL oder 125 g nicht mit H und P S tzen gekennzeichnet werden Component Hazard contents GHS symbol Hazard Precaution phrases phrases Inhalt Gefahrstoff GHS Symbol H S tze P S tze A2 Sodium hydroxide 0 5 2 96 Warning 290 315 234 280 302 352 Natriumhydroxid 0 5 2 Achtung 319 305 351 338 332 313 337 313 390 406 A3 Guanidine hydrochloride 0 Warning 302 319 280 301 312 Guanidinhydrochlorid Achtung 305 351 338 330 337 313 RNase A RNase A Danger 317 334 261 3024352 RNase A lt gt Gefahr 304 340 333 313 342 311 363 Hazard phrases H 290 May be corrosive to metals Kann gegen ber Metallen korrosiv sein H 302 Harmful if swallowed Gesundheitssch dlich bei Verschlucken H 315 Causes skin irritation Verursacht Hautreizungen H 317 May cause an allergic skin reaction Kann allergische Hautreaktionen verursachen H 319 Causes serious eye irritation Verursacht schwere Augenreizung H 334 May cause allergy or asthma symptoms or breathing difficulties if inhaled Kann bei Einatmen Allergie asthmaartige Symptome oder Atembeschwerden verursa chen Precaution phrases P 234 Keep only in original container Nur im Originalbeh lter aufbewahren P 261 Avoid breathing dust Einatmen von Staub vermeiden P 280 Wear protective gloves eye protection Schutzhandschuhe Augenschutz tragen MACHEREY
20. tive compounds when combined with bleach sodium hypochlorite DO NOT add bleach or acidic solutions directly to the sample preparation waste All kit components can be stored at room temperature 18 25 C and are stable for at least one year Always keep buffer bottles tightly closed especially if buffers are preheated during the preparation Storage of Buffer A2 below 20 C may cause precipitation of SDS If salt precipitate is observed incubate buffer at 30 40 C for several minutes and mix well until all precipitate is redissolved completely Cool down to room temperature before use Before starting any NucleoSpin Plasmid EasyPure protocol prepare the following Add Liquid RNase A to Buffer A1 and mix thoroughly Indicate date of RNase A addition Store Buffer A1 containing RNase A at 4 C The solution will be stable at this temperature for at least six months Add the indicated volume of 96 100 ethanol to Buffer AQ Concentrate NucleoSpin Plasmid EasyPure 10 preps 50 preps 250 preps 740727 10 740727 50 740727 250 6mL 6mL 25mL Add 24 mL ethanol Add 24 mL ethanol Add 100 mL ethanol Wash Buffer AQ Concentrate 10 MACHEREY NAGEL 07 2014 Rev 02 Plasmid DNA purification 4 Safety instructions The following components of the NucleoSpin Plasmid EasyPure kit contain hazardous contents GHS classification Only harmful features do not need to be labeled with H and P ph
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