Hongo 3

w
e
b
BIO 101 Protocol
RPM
Rapid Pure Miniprep Kit
2070-000
2070-200
2070-400
2070-500
2070-600
6
60
120
300
600
preps
preps
preps
preps
preps
Rapid Isolation and Purification of DoubleStranded Plasmid DNA from Bacterial Cultures
12 preps in less than 12 Minutes
GENECLEAN-Quality Pure Plasmid DNA for
Cutting, Cloning, Sequencing, Amplification
No Phenol/Chloroform Extractions
Shipping & Storage:
The RPM Kit is shipped and stored at ambient temperature.
Check with BIO 101...
We have you time in mind!
Revision #: 2070-999-7C03P
1070 Joshua Way, Vista CA 92083, USA

Tel: (760) 598-7299 Fax (760)598-0116
E-mail: technical@bio101.com
Website: www.bio101.com
For technical support call (800) 424-6101
RPM Protocol
Kit Components: .......................................................................... 3
2070-000 (6 preps).................................................................. 3
2070-200 (60 preps) ...............................................................3
2070-400 (120 preps) .............................................................4
2070-500 (300 preps) .............................................................4
2070-600 (600 preps) .............................................................5
Introduction ................................................................................. 6
Protocol ....................................................................................... 7
Notes ........................................................................................... 9
Product Use Limitation & Warranty .......................................... 10
General Information .............................................................10
Call (800) 424-6101 for Technical Support
Kit Components:
2070-000 (6 preps)
Name
Volume
Cat #
Pre-Lysis Buffer
1.0 ml
Alkaline Lysis Solution
1.0 ml
Neutralizing Solution
1.0 ml
GLASSMILK SpinBuffer*
1.8 ml
Wash Solution Concentrate**
2.0 ml
SPIN Filters
6
Catch Tubes
6
CIRCLEGROW Sample
8 capsules
2070-001
2070-002
2070-003
2070-004
2070-005
2070-006
2070-007
3000-101
*GLASSMILK SpinBuffer contains guanidine thiocyanate, use with

proper precaution.
**Add 2.0 ml of 100% Ethanol to Wash Solution Concentrate and
mix before initial use.
2070-200 (60 preps)

Name
Volume
Cat #
Pre-Lysis Buffer
4.0 ml
7.0 ml
7.0 ml
16 ml
12 ml
SPIN Filters
60
Catch Tubes
60
CIRCLEGROW Sample
8 capsules
2070-201
2070-202
2070-203
2070-204
2070-205
2070-206
2070-207
3000-101

proper precaution.
**Add 12 ml of 100% Ethanol to Wash Solution Concentrate and mix
before initial use.
Call (800) 424-6101 for Ordering Information
2070-400 (120 preps)

Name
Volume
Cat #
Pre-Lysis Buffer
7.0 ml
15 ml
15 ml
35 ml
24 ml
SPIN Filters
120
Catch Tubes
120
CIRCLEGROW Sample 20 capsules
2070-401
2070-402
2070-403
2070-404
2070-405
2070-406
2070-407
3000-011
*GLASSMILK Spinbuffer contains guanidine thiocyanate, use with

proper precaution.
**Add 24 ml of 100% Ethanol to Wash Solution Concentrate and
2070-500 (300 preps)

Name
Volume
Cat #
Pre-Lysis Buffer
18 ml
33 ml
33 ml
90 ml
60 ml
SPIN Filters
300
Catch Tubes
300
2070-501
2070-502
2070-503
2070-504
2070-505
2070-506
2070-507
3000-011
*GLASSMILK Spinbuffer contains guanidine thiocyanate, use with

proper precaution.
**Add 60ml of 100% Ethanol to Wash Solution Concentrate and mix
before initial use.
2070-600 (600 preps)

Name
Volume
Cat #
Pre-Lysis Buffer
35 ml
65 ml
65 ml
180 ml
120 ml
SPIN Filters
5 x 120
Catch Tubes
5 x 120
2070-601
2070-602
2070-603
2070-604
2070-605
2070-606
2070-607
3000-001

proper precaution.
**Add 120ml of 100% Ethanol to Wash Solution Concentrate and
All references to user supplied reagents are italicized and

bolded in the protocol for your convenience.
Introduction
The RPM Kit delivers up to 20 g of supercoiled plasmid DNA with
minimum effort and time. Multiple samples can be processed rapidly by
using 1.5 ml of bacterial culture. With practice, the procedure can be
completed in less than 12 minutes, yielding pure plasmid DNA suitable for
enzymatic manipulations including DNA sequencing, restriction enzyme
digestion, in vitro transcription1, and PCR2. Reagents and materials supplied
by the user are 100% ethanol, sterile H2O or TE buffer, 1.5 ml microcentrifuge
vials (one for each miniprep), and a microcentrifuge. The reagents and materials
provided with the kit are for research use only.
1
2
For use of the plasmid DNA for in vitro transcription experiments, addition of RNase
Inhibitor is strongly recommended.
PCR Process is covered by U.S. Patent Numbers 4.683.202 and 4.683.195 issued to
Hoffman & LaRoche Corporation.
Protocol
1.
Spin 1.5 ml of bacterial culture (use Circlegrow media to grow cells

to high density for maximum yield of plasmid DNA) in a
microcentrifuge vial for 30 seconds, remove and discard supernatant. If
use of more than 1.5 ml culture for this miniprep is desired, add more
culture, re-spin, remove and discard supernatant. (Pour off supernatant
and then remove the remaining few microliters with a pipet tip.)
2.
To resuspend the cell pellet, add 50 l Pre-Lysis Buffer and mix by

vortexing or by pipetting up and down until completely suspended.
3.
Add 100 l Alkaline Lysis Solution directly into the cell suspension
and mix by inverting the tube several times. Before proceeding to
step 4, wait 1 minute to allow lysis to complete (solution should be
clear).
This is a critical step in the procedure and affects yield and purity
of the plasmid DNA; the cells must be homogeneously lysed and
some host cells lyse easier than others. For optimum consistency of
lysis from one cell type to another, be certain the Alkaline Lysis
Solution is 22-25C and that the SDS is in solution.
4.
Add 100 l Neutralizing Solution and mix by briefly vortexing. A

white precipitate will form consisting of cell membranes, proteins, and
chromosomal DNA. Spin in a microcentrifuge for 2 minutes to pellet
white precipitate.
4a. Prepare a SPIN Filter for each sample by adding 250 l

GLASSMILK SpinBuffer (mix suspension completely before
each use) to the SPIN Filter chamber and placing it in a
Catch Tube.
4b. Remove supernatant from sample while avoiding white pellet,
and transfer to a prepared SPIN Filter (see step 4a).
8
5.
Pipet up and down a couple of times to mix and, after 1 minute, spin
for 1 minute to collect liquid in the bottom of the Catch Tube.
6.
Add 350 l Wash Solution (remember to add an equal volume of

ethanol before first use)* to SPIN Filter and spin for 1 minute. Pour
off liquid in collection vial and spin again for 1 minute to drive last of
the liquid out of the SPIN Filter. Transfer SPIN Filter to a new
Catch Tube. Discard other tube.
7.
To elute plasmid DNA from the SPIN Filter, add 50 l of autoclaved

sterile H2O or TE buffer (10 mM Tris-HCl, pH 7.5, 1 mM EDTA),
vortex briefly (no more than half speed) to resuspend GLASSMILK, and
spin for 30 seconds to collect DNA in the bottom of the Catch Tube.
Remove Catch Tube from microcentrifuge; remove and discard
SPIN Filter. Close Catch Tube containing plasmid DNA and store
at +4C or -20C. Note: The yield of plasmid DNA is dependent on the
bacterial host and the plasmid copy number. The plasmid DNA is now
ready for further manipulation.
*
Catalog #
Description
Ethanol
to add*
Wash Sol.
Conc.
Final
Volume
2070-005
RPM Sample
2 ml
2 ml
4 ml
2070-205
RPM-60
12 ml
12 ml
24 ml
2070-405
RPM-120
24 ml
24 ml
48 ml
2070-505
RPM-300
60 ml
60 ml
120 ml
2070-605
RPM-600
120 ml
120 ml
240 ml
* 100% Ethanol is provided by the user.
Notes
10
Notes
11
Notes
Product Use Limitation & Warranty

Unless otherwise indicated, this product is for research use only. Purchase
of BIO 101 products does not grant rights to reproduce, modify, repackage the
products or any derivative thereof to third parties. BIO 101 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 BIO 101's sole liability
hereunder shall be limited to, at our option, product credits, refund of the
purchase price of, or the replacement of all material(s) that does not meet our
specification. By acceptance of the product, Buyer indemnifies and holds BIO
101 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 refunds or replacement is conditioned on Buyer notifying
BIO 101 within thirty (30) days of receipt of product. Failure of Buyer to give
said notice within thirty (days) shall constitute a waiver by the Buyer of all
claims hereunder with respect to said material(s).
General Information
BIO 101 is a pioneer in developing kits for molecular biology research. We
introduced the GENECLEAN Kits in 1986 and have since been manufacturing
products to bring convenience into your research. Our goal is to make your life
easier by simplifying the complexities of lab work.
Technical Support and Ordering Information
(800) 424-6101 (Toll Free - Continental USA)
(760) 598-7299 (Outside USA)
(760) 598-0116 (Fax line)
technical@bio101.com
order@bio101.com
http://www.bio101.com
Office Hours:
6:30 am - 6:00 pm P.S.T. (Mon - Fri)
Mailing Address:
BIO 101, Inc.
P.O. Box 2284
La Jolla, CA 92038-2284 USA

Hongo 3

Uploaded by

Copyright:

Available Formats

Hongo 3

Uploaded by

Document Information

Original Description:

Original Title

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Hongo 3

Uploaded by

Copyright:

Available Formats

w

BIO 101 Protocol

1070 Joshua Way, Vista CA 92083, USA

For technical support call (800) 424-6101

Call (800) 424-6101 for Technical Support

*GLASSMILK SpinBuffer contains guanidine thiocyanate, use with

2070-200 (60 preps)

*GLASSMILK SpinBuffer contains guanidine thiocyanate, use with

Call (800) 424-6101 for Ordering Information

2070-400 (120 preps)

*GLASSMILK Spinbuffer contains guanidine thiocyanate, use with

2070-500 (300 preps)

*GLASSMILK Spinbuffer contains guanidine thiocyanate, use with

Call (800) 424-6101 for Technical Support

2070-600 (600 preps)

*GLASSMILK SpinBuffer contains guanidine thiocyanate, use with

All references to user supplied reagents are italicized and

Call (800) 424-6101 for Ordering Information

Call (800) 424-6101 for Technical Support

Spin 1.5 ml of bacterial culture (use Circlegrow media to grow cells

To resuspend the cell pellet, add 50 l Pre-Lysis Buffer and mix by

Add 100 l Neutralizing Solution and mix by briefly vortexing. A

4a. Prepare a SPIN Filter for each sample by adding 250 l

Call (800) 424-6101 for Ordering Information

Add 350 l Wash Solution (remember to add an equal volume of

To elute plasmid DNA from the SPIN Filter, add 50 l of autoclaved

* 100% Ethanol is provided by the user.

Call (800) 424-6101 for Technical Support

Call (800) 424-6101 for Ordering Information

Call (800) 424-6101 for Technical Support

Call (800) 424-6101 for Ordering Information

Product Use Limitation & Warranty

You might also like