INSTRUCTIONAL MANUAL
NEWCCA Plasmid Miniprep Kits
Catalog Numbers: TK-EP0050, TK-EP0100
For purification of high quality plasmid DNA for molecular biology applications
Patent Pending; For Research Use Only; Not for Diagnosis or Therapy Purposes.
©NEWCCA, 2111 Crescent Ave., Suite G, Anaheim, California 92801, USA
Tel: 1-310-666-1999; Fax: 1-310-496-6150
Website: www.newcca.com
email: sales@newcca.com
Page 1 of 6
Components
NEWCCA Plasmid
Miniprep Kit
P1 buffer
P2 buffer1
P3 buffer1
Wash buffer2
Elution buffer3
RNase A1 (10mg/ml)
Spin column-1
Collection Tube
Manual
TK-EP0050
(50 preps)
12 mL
12 mL
18 mL
8 mL
4 mL
200 µL
50
50
1
TK-EP0100
(100 preps)
22 mL
22 mL
35 mL
16 mL
6 mL
400 µL
100
100
1
Storage Temperature
4 °C after adding RNase A
Room Temp.
Room Temp.
Room Temp.
Room Temp.
-20 °C upon receiving
Room Temp.
Room Temp.
--
Note: Integrity of kit components is guaranteed for up to one year from date of purchase.
Reagents are routinely tested on a lot-to-lot basis to ensure they provide maximal performance
and reliability.
1
Caution:
P2 Buffer contains NaOH: irritant and corrosive. Risk and safety phrases: R35, (S1/2), S26,
S37/39, S45.
P3 Buffer contains chaotropic reagents: Harmful. Risk and safety phrases: R20/21/22-32, S1326-36-46.
RNase A contains ribonuclease: sensitizer. Risk and safety phrase: R42/43, S23-24-26-36/37.
Please use proper safety precautions with these reagents.
2
Add ethanol to Wash Buffer (concentrate) prior to use. See Buffer Preparation for
instructions.
3
Elution buffer: 10 mM Tris-HCl, pH 8.0, 1 mM EDTA
Buffer Preparation
1. Add ethanol to the wash buffer at a 4:1 volume ratio of ethanol to buffer.
For products TK-EP-0050, add 32 mL of 100% ethanol or 42.6 mL 95% ethanol to each 8 mL
Wash Buffer. For products TK-EP-0100, add 64 mL of 100% ethanol or 85.2 mL 95% ethanol to
each 16 mL Wash Buffer.
2. Add 200 µL (TK-EP0050) or 400 µL (TK-EP0100) RNase A into P1 Buffer, and store at 2-8 °C.
©NEWCCA, 2111 Crescent Ave., Suite G, Anaheim, California 92801, USA
Tel: 1-310-666-1999; Fax: 1-310-496-6150
Website: www.newcca.com
email: sales@newcca.com
Page 2 of 6
Introduction
NEWCCA Plasmid Miniprep Kits are designed for efficient extract of high quality plasmid DNA from E.
Coli in a very simple, rapid, reliable and cost-effective procedure. NEWCCA Plasmid Miniprep Kits use
silica membrane technology to eliminate phenol extraction/ethanol precipitation, and other steps
associated with loose resins or slurries. The plasmid DNA purified from NEWCCA Plasmid Miniprep Kits
are suitable for various downstream applications, including restriction enzyme digestion, sequencing,
ligation, transfection, transformation, cloning, PCR, in vivo injection, etc.
NEWCCA Plasmid Miniprep Kits procedure is based on silica membrane technology. The crude plasmid
DNA is released from bacterial cells by an alkaline lysis and absorbed into the silica membrane. The
contaminations in crude plasmid DNA such as proteins and endotoxins will be washed away by washing
steps. Finally the plasmid DNA is eluted by water or Elution Buffer.
Four basic steps are involved in this procedure:
 Preparation of a bacterial lysate containing plasmid DNA.
 Binding of plasmid DNA into NEWCCA Spin Column-1;
 Washing away of impurities;
 Elution of plasmid DNA.
Specifications
 DNA purity: Typical Abs260/280 ≥ 1.8; Endotoxin-free.
 DNA yield: Typical 25 µg from 5 mL culture per column, depending on the plasmid copy number,
bacterial culture conditions and strains, etc.

DNA size: Up to 25 kb.

Elution volume: 50 µL.

Cost efficiency.
©NEWCCA, 2111 Crescent Ave., Suite G, Anaheim, California 92801, USA
Tel: 1-310-666-1999; Fax: 1-310-496-6150
Website: www.newcca.com
email: sales@newcca.com
Page 3 of 6
Procedure Outline
©NEWCCA, 2111 Crescent Ave., Suite G, Anaheim, California 92801, USA
Tel: 1-310-666-1999; Fax: 1-310-496-6150
Website: www.newcca.com
email: sales@newcca.com
Page 4 of 6
Procedure
1. Centrifuge 1 - 10 mL of bacterial culture at 5,000 rpm for 5 minutes.
2. Discard supernatant.
3. Add 200 μL of P1 Buffer (cold) to the tube and resuspend pellet completely (i.e., by
vortexing or pipeting), and transfer to a 1.5-mL microcentrifuge tube.
4. Add 200 μL of P2 Buffer and mix thoroughly by inverting the tube 4 - 5 times. Incubate at room
temperature for 3-4 minutes.
5. Add 300 μL of P3 Buffer and mix gently but thoroughly by inverting the tube 4 - 5 times. Do not
vortex. Allow the lysate to incubate on ice for 10 minutes.
6. Centrifuge sample(s) for 10 minutes at ≥10,000 x g in a table-top microcentrifuge at 4 °C.
7. Place a Spin Column-1 in a Collection Tube and transfer the supernatant from Step 6 into the
Spin Column-1. Be careful not to disturb the pellet to avoid transferring any cellular debris to the
column.
8. Centrifuge the Spin Column-1/Collection Tube assembly at ≥10,000 x g for 30 seconds.
9. Discard the flow-through in the Collection Tube, making sure the flow-through does not touch
the bottom of the column. Return the Spin Column-1 to the Collection Tube.
10. Add 500 μL of Wash Buffer to the column and centrifuge at ≥10,000 x g for 30 seconds.
11. Discard the flow-through in the Collection Tube, making sure the flow-through does not touch
the bottom of the column. Return the Spin Column-1 to the Collection Tube.
12. Add 250 μL of Wash Buffer to the column and centrifuge at ≥10,000 x g for 1 minute.
13. Empty the Collection Tube, centrifuge Spin Column-1/Collection Tube assembly at ≥10,000 x
g for 1 minute.
14. Transfer the Spin Column-1 into a clean 1.5-mL microcentrifuge tube and then add 50 μL of
Elution Buffer to the center of Spin Column-1. Let stand for 1 minute, and centrifuge at
≥10,000 x g for 1 minute to elute the plasmid DNA.
©NEWCCA, 2111 Crescent Ave., Suite G, Anaheim, California 92801, USA
Tel: 1-310-666-1999; Fax: 1-310-496-6150
Website: www.newcca.com
email: sales@newcca.com
Page 5 of 6
Ordering Information
Product
Contents Cat. No.
Plasmid DNA Miniprep Kits
50 preps.
100 preps.
50 preps.
100 preps.
50 mL
100 mL
50 mL
100 mL
50 mL
100 mL
50 mL
100 mL
8 mL
16 mL
10 mL
50 mL
50
100
50
100
Gel Recovery/Clean-up All-in-One™ Kits
P1 Buffer
P2 Buffer
P3 Buffer
All-in-One™ (AOB) Buffer
Wash Buffer (Concentrate)
Elution Buffer
NEWCCA Spin Column-1 with Collection Tube
NEWCCA Spin Column-2 with Collection Tube
TK-EP0050
TK-EP0100
TK-GR0050
TK-GR0100
TK-EP-P1-0050
TK-EP-P1-0100
TK-EP-P2-0050
TK-EP-P2-0100
TK-EP-P3-0050
TK-EP-P3-0100
TK-GR-AOB-0050
TK-GR-AOB-0100
TK-WB-008
TK-WB-006
TK-EB-010
TK-EB-050
TK-SC1-050
TK- SC1-100
TK-SC2-050
TK- SC2-100
©NEWCCA, 2111 Crescent Ave., Suite G, Anaheim, California 92801, USA
Tel: 1-310-666-1999; Fax: 1-310-496-6150
Website: www.newcca.com
email: sales@newcca.com
Page 6 of 6