Macherey-Nagel NucleoSpin 740727.10 User manual

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Plasmid DNA
purication
User manual
NucleoSpin®Plasmid EasyPure
August 2013 / Rev. 01
A0xxxxx /01308

Plasmid DNA purication
Protocol-at-a-glance (Rev. 01)
MACHEREY-NAGEL GmbH & Co. KG · Neumann-Neander-Str. 6–8 · 52355 Düren · Germany
Tel.: [phone] · Fax: [phone] · [email protected] · www.mn-net.com
NucleoSpin®
Plasmid EasyPure
1Cultivate
and harvest bacterial
cells
12,000 x g,
30 s
2Cell lysis 150 μL Buffer A1
250 μL Buffer A2
RT, up to 2 min
350 μL Buffer A3
3Clarication of the lysate
> 12,000 x g,
3 min
4Bind DNA Load supernatant
1,000–2,000 x g,
30 s
5Wash and dry silica
membrane 450 μL Buffer AQ
> 12,000 x g,
1 min
6 Elute DNA 50 μL Buffer AE
RT, 1 min
> 12,000 x g,
1 min

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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specications 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
4.1 Risk and safety phrases 11
4.2 GHSclassication 12
5 NucleoSpin®Plasmid EasyPure protocol 14
6 Appendix 16
6.1 Troubleshooting 16
6.2 Ordering information 19
6.3 References 19
6.4 Product use restriction / warranty 20

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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 5 mL 15 mL 75 mL
Lysis Buffer A2 5 mL 15 mL 75 mL
Neutralization Buffer A3 5 mL 20 mL 100 mL
Wash Buffer AQ
(Concentrate)* 6 mL 6 mL 30 mL
Elution Buffer AE** 15 mL 15 mL 30 mL
Liquid RNase A 2 mg 6 mg 30 mg
NucleoSpin®Plasmid
EasyPure Columns
(dark blue rings)
10 50 250
Collection Tubes (2 mL) 10 50 250
Short protocol 111
* For preparation of working solutions and storage conditions see section 3.
**Composition of Elution Buffer AE: 5 mM Tris/HCl, pH 8.5

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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
• 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rst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purication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.

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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nallyelutedunderlowionic
strength conditions with slightly alkaline Buffer AE (5 mM Tris/HCl, pH 8.5).
2.2 Kitspecications
• The NucleoSpin®Plasmid EasyPure kits are designed for the rapid, small-
scale preparation of highly pure plasmid DNA (mini preps).
• The NucleoSpin®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puriedplasmidDNAissuitableforapplicationslikeautomateduorescent
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

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Table1:Kitspecicationsataglance
Parameter NucleoSpin®Plasmid EasyPure
Culture volume 2–10 mL
Typical yield 15–30μg
Elution volume 50μL
Binding capacity 35μg
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. Usually an
OD of 3–6canbeachieved.Alternatively,richmedialike2 xYT(Yeast / Tryptone),TB
(TerricBroth),orCircleGrowcanbeused.Inthiscasebacteriagrowfaster,reachthe
stationaryphasemuchearlierthaninLBmedium(≤ 12 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
nd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

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Table 2: Information about antibiotics according to Maniatis*
Antibiotic Stock solution
(concentration)
Storage Working
concentration
Ampicillin 50 mg/mL in H2O-20 °C 20–50μg/mL
Carbenicillin 50 mg/mL in H2O-20 °C 20–60μg/mL
Chloramphenicol 34 mg/mL in EtOH -20 °C 25–170μg/mL
Kanamycin 10 mg/mL in H2O-20 °C 10–50μg/mL
Streptomycin 10 mg/mL in H2O-20 °C 10–50μg/mL
Tetracycline 5 mg/mL in EtOH -20 °C 10–50μg/mL
As rule of thumb use 5 mLofawellgrownLBcultureasgiveninthekitspecications.
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 (OD600).
Table 3: Recommended culture volumes for NucleoSpin®Plasmid EasyPure
OD600 123456
Culture volume 15 mL 8 mL 5 mL 4 mL 3 mL 2 mL
Note, if too much bacterial material is used, the lysis and precipitation steps become
inefcient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μL 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 cloning. A laboratory manual, Cold Spring Harbor, Cold Spring,
New York 1982.

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• 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 %). 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 CO2leads to a decrease in
pH of unbuffered solutions.

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10
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 reactive
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
REF 740727.10 740727.50 740727.250
Wash Buffer AQ
(Concentrate)
6 mL
Add 24 mL ethanol
6 mL
Add 24 mL ethanol
30 mL
Add 120 mL ethanol

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4 Safety instructions
The following components of the NucleoSpin®Plasmid EasyPure kit contain
hazardous contents.
4.1 Risk and safety phrases
Component Hazard contents Hazard
symbol
Risk
phrases
Safety
phrases
Inhalt Gefahrstoff Gefahrstoff-
symbol
R-Sätze S-Sätze
A2 Sodium hydroxide 0.5–2 % Xi* R 36/38 S 26-
37/39-45
Natriumhydroxid 0.5–2 %
A3 Guanidine hydrochloride 36–50 % Xn** R 22-36 S 26-39
Guanidinhydrochlorid 36–50 %
RNase A RNase A Xn R 42/43 S 22-24
RNase A
Risk phrases
R 22 Harmful if swallowed. R 36 Irritating to eyes.
Gesundheitsschädlich beim Verschlucken. Reizt die Augen.
R 36/38 Irritating to eyes and skin. R 42/43 May cause sensitization by inhala-
tion and skin contact.
Reizt die Augen und die Haut.
Sensibilisierung durch Einatmen und
Hautkontakt möglich.
Safety phrases
S 22 Do not breathe dust.
Staub nicht einatmen.
S 24 Avoid contact with the skin.
Berührung mit der Haut vermeiden.
S 26 In case of contact with eyes, rinse immediately with plenty of water and seek
medical advice.
Bei Berührung mit den Augen gründlich mit Wasser abspülen und Arzt konsultieren.
S 37/39 Wear suitable gloves and eye / face protection.
Bei der Arbeit geeignete Schutzhandschuhe und Schutzbrille / Gesichtsschutz tragen.
S 39 Wear suitable eye / face protection.
Schutzbrille / Gesichtsschutz tragen.
S 45 In case of accident or if you feel unwell, seek medical advice immediately (show
the label where possible).
Bei Unfall oder Unwohlsein sofort Arzt hinzuziehen (wenn möglich, dieses Etikett vorzeigen).
* Hazard labeling not neccessary if quantity per bottle below 25g or mL (certicate of exemption
according to 67/548/EEC Art. 25, 1999/45/EC Art. 12 and German GefStoffV § 20 (3) and TRGS 200 7.1).
For further information see Material Safety Data Sheet.
**Hazard labeling not neccessary if quantity per bottle below 125g or mL (certicate of exemption
according to 67/548/EEC Art. 25, 1999/45/EC Art. 12 and German GefStoffV § 20 (3) and TRGS 200 7.1).
For further information see Material Safety Data Sheet.

MACHEREY-NAGEL – 08 / 2013, Rev. 01
12
4.2 GHSclassication
Only harmful features do not need to be labeled with H and P phrases until 125 mL or
125 g.
Mindergefährliche Eigenschaften müssen bis 125 mL oder 125 g nicht mit H- und P-Sätzen gekennzeichnet
werden.
Component Hazard contents GHS symbol Hazard
phrases
Precaution
phrases
Inhalt Gefahrstoff GHS Symbol H-Sätze P-Sätze
A2 Sodium hydroxide 0.5–2 % Warning 290, 315,
319
234, 280, 302+352,
305+351+338,
332+313, 337+313,
390, 406
Natriumhydroxid 0.5–2 % Achtung
A3 Guanidine hydrochloride Warning 302, 319 280, 301+312,
305+351+338, 330,
337+313
Guanidinhydrochlorid Achtung
RNase A RNase A Danger 317, 334 261, 302+352,
304+341, 333+313,
342+311, 363
RNase A Gefahr
Hazard phrases / H-Sätze
H 290 May be corrosive to metals.
Kann gegenüber Metallen korrosiv sein.
H 302 Harmful if swallowed.
Gesundheitsschädlich 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difcultiesifinhaled.
Kann bei Einatmen Allergie, asthmaartige Symptome oder Atembeschwerden verursa-
chen.
Precaution phrases / P-Sätze
P 234 Keep only in original container.
Nur im Originalbehälter aufbewahren.
P 261 Avoid breathing dust.
Einatmen von Staub vermeiden.
P 280 Wear protective gloves / eye protection.
Schutzhandschuhe /Augenschutz tragen.
P 301+312 IF SWALLOWED: Call a POISON CENTER or doctor /physician if you feel
unwell.
BEI VERSCHLUCKEN: Bei Unwohlsein GIFTINFORMATIONSZENTRUM oder Arzt
anrufen.
P 302+352 IF ON SKIN: Wash with plenty of soap and water.
BEI KONTAKT MIT DER HAUT: Mit viel Wasser und Seife waschen.
PlasmidDNApurication

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MACHEREY-NAGEL – 08/ 2013, Rev. 01
Precaution phrases / P-Sätze
P 304+341 IFINHALED:Ifbreathingisdifcult,removevictimtofreshairandleepat
rest in a position comfortable for breathing.
BEI EINATMEN: Bei Atembeschwerden an die frische Luft bringen und in einer Position
ruhigstellen, die das Atmen erleichtert.
P 305+351+338 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 spülen.
Vorhandene Kontaktlinsen nach Möglichkeit entfernen. Weiter spülen.
P 330 Rinse mouth.
Mund ausspülen.
P 332+313 If skin irritation occurs: Get medical advice / attention.
Bei Hautreizung: Ärztlichen Rat einholen / ärztliche Hilfe hinzuziehen.
P 333+313 IF skin irritation or a rash occurs: Get medical advice / attention.
Bei Hautreizung oder -ausschlag: Ärztlichen Rat einholen / ärztliche Hilfe hinzuziehen.
P 337+313 Get medical advice / attention.
Bei anhaltender Hautreizung: Ärztlichen Rat einholen / ärztliche Hilfe hinzuziehen.
P 342+311 If experiencing respiratory symptoms: Call a POISON CENTER or doc-
tor / physician.
Bei Symptomen der Atemwege: GIFTINFORMATIONSZENTRUM oder Arzt anrufen.
P 363 Wash contaminated clothing before reuse.
Kontaminierte Kleidung vor erneutem Tragen waschen.
P 390 Absorb spillage to prevent material damage.
Verschüttete Menge aufnehmen, um Materialschäden zu vermeiden.
P 406 Store in a corrosive resistant container with a resistant inner liner.
In korrosionsbeständigem Behälter mit korrosionsbeständiger Auskleidung aufbewahren.
For further information please see Material Safety Data Sheets (www.mn-net.com).
Weiterführende Informationen nden Sie in den Sicherheitsdatenblättern (www.mn-net.com).
PlasmidDNApurication

MACHEREY-NAGEL – 08 / 2013, Rev. 01
14
5 NucleoSpin®Plasmid EasyPure protocol
Before starting the preparation:
• Check if Wash Buffer AQ was prepared according to section 3.
1 Cultivate and harvest bacterial cells
Use 2–10 mL 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.
Discard the supernatant and remove as much of the
liquid as possible.
> 12,000 x g,
30 s
2Cell lysis
Add 150μL BufferA1. Resuspend the cell pellet
completely by vortexing or pipetting up and down. Make
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
at 30–40 °C until precipitate is dissolved completely. Cool
buffer down to room temperature (18–25 °C) before use.
Add 250μL BufferA2. Mix gently by inverting the tube
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μL BufferA3. Mix thoroughly by inverting the
tube until LyseControl has turned colorless throughout
the lysate without any traces of blue color. Do not vortex
to avoid shearing of genomic DNA!
+ 150 μL A1
Resuspend
+ 250 μL A2
Mix
RT, 2 min
+ 350 μL A3
Mix
3Claricationoflysate
Centrifuge for 3 min at full speed (> 12,000 x g).
> 12,000 x g,
3 min
NucleoSpin®Plasmid EasyPure

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MACHEREY-NAGEL – 08/ 2013, Rev. 01
4 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ow-throughandplacethespincolumnbackinto
the collection tube.
Load
supernatant
1,000–
2,000 x g,
30 s
5Wash and dry silica membrane
Add 450μLBufferAQ (supplemented with ethanol, see
section 3).
Centrifuge for 1 min at full speed (> 12,000 x g).
Very carefully discard the collection tube and the ow-
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 °C to dry silica membrane completely.
+ 450 μLAQ
> 12,000 x g,
1 min
6Elute DNA
Place the NucleoSpin®Plasmid EasyPure Column in
a 1.5 mL microcentrifuge tube (not provided) and add
50μLBufferAE.
Incubate for 1 min at room temperature (18–25 °C).
Centrifuge for 1 min at full speed (> 12,000 x g).
Note: For more efcient elution procedures and alternative
elution buffer (e.g., TE buffer or water) see section 2.4.
+ 50 μL AE
RT, 1 min
> 12,000 x g,
1 min
NucleoSpin®Plasmid EasyPure

MACHEREY-NAGEL – 08 / 2013, Rev. 01
16
6 Appendix
6.1 Troubleshooting
Problem Possible cause and suggestions
Incomplete
lysis of
bacterial
cells
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.
SDS in Buffer A2 precipitated
• 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.
Too many bacterial cells used
• See table 3 for maximum amount of cells.
Poor
plasmid
yield
Incomplete lysis of bacterial cells
• See„Possiblecauseandsuggestions“above.
No or insufcient amounts of antibiotic used during cultivation
• Cells carrying the plasmid of interest may become overgrown
by cells without plasmid (see table 2), when inadequate levels
of the antibiotics are used. Add appropriate amounts of freshly
prepared stock solutions to all media; both solid and liquid.
Bacterial culture too old
• Do not incubate cultures for more than 16 h (LB) or 12 h
(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.
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Poor
plasmid
yield
(continued)
Suboptimal elution conditions
• If 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
pHofthewater.Elutionefcienciesdropdrasticallywithbuffers
< pH 7.
Low copy-number plasmid was used
• At least double or triple culture volume and increase lysis buffers
if nal amount of cells exceed the recommended volumes of
table 3.
No plasmid
yield
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
• Especially when working with nuclease-rich strains, keep
plasmid preparations on ice or frozen in order to avoid DNA
degradation.
Inappropriate storage of plasmid DNA
• Quantitate DNA directly after preparation, for example, by
agarose gel electrophoresis. Store plasmid DNA dissolved in
water at < -18 °C or at < +5 °C when dissolved in Buffer AE or
TE buffer.
Poor
plasmid
quality
Nicked plasmid DNA
• Cell suspension was incubated with alkaline Lysis Buffer A2 too
long. Reduce lysis time.
Genomic DNA contamination
• Cell lysate was vortexed or mixed too vigorously after addition
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.
PlasmidDNApurication

MACHEREY-NAGEL – 08 / 2013, Rev. 01
18
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ow-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.
PlasmidDNApurication

19
MACHEREY-NAGEL – 08/ 2013, Rev. 01
6.2 Ordering information
Product REF Pack of
NucleoSpin®Plasmid EasyPure 740727.10 / .50 / .250 10 / 50 / 250preps
NucleoSpin®Buffer Set
(for the isolation of low-copy plasmids) 740953 1
Buffer A1
(without RNase A) 740911.1 1 L
Buffer A2 740912.1 1 L
Buffer A3 740913.1 1 L
Buffer AQ Concentrate
(for 100 mL Buffer AQ) 740995 20 mL
Buffer AE 740917.1 1 L
Liquid RNase A 740397 250 mg
Collection Tubes (2 mL) 740600 1000
6.3 References
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puricationofDNA
from agarose. Proc. Natl. Acad. Sci. USA 76: 615 - 619.
PlasmidDNApurication

MACHEREY-NAGEL – 08 / 2013, Rev. 01
20
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leaets.
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 elds of application.Application on the human body is STRICTLY
FORBIDDEN. The respective user is liable for any and all damages resulting from such
application.
DNA/RNA/PROTEIN purication 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 specic 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puricationproductsofMACHEREY-NAGELforawell-denedand
specicapplication.
MACHEREY-NAGELshallonlyberesponsiblefortheproductspecicationsandthe
performancerangeofMNproductsaccordingtothespecicationsofin-housequality
control, product documentation and marketing material.
ThisMACHEREY-NAGELproductisshippedwithdocumentationstatingspecications
and other technical information. MACHEREY-NAGEL warrants to meet the stated
specications.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
PlasmidDNApurication