TM
EasyPrep
DNA/RNA
Miniprep Manual
Table of Contents
Introduction.....................................................
3
Kit Contents ...................................................
4
Specifications……………………….………..
4

For simultaneous purification of genomic DNA and
RNA from the same sample
Buffer Preparation............................................ 4

Catalog#: DR01-01, DR01-02
Storage and Stability......................................
5
DNA/RNA Purification Protocols
Tissues…………………………...……... 6
Cells…..………………………...……...
8
Whole blood, serum and plasma…….
9
Trouble Shooting Guide .................................
10
Other Related Products……………………….. 11
For research use only
(November 2013)
2
EasyPrepTM DNA/RNA Miniprep
Introduction
The EasyPrepTM DNA/RNA Kit provides a simple and rapid method for
simultaneously isolating genomic DNA and total RNA from a biological
samples, which includes cultured cells, tissues, whole blood, plasma,
serum, and body fluids. This co-purification system saves time on sample
handling, avoids the sample variation and saves precious samples, which
is essential when the amount of available samples are limited
Instead of using proteinase K to
digest the biological materials, the
EasyPrepTM DNA/RNA Kit applies
the lysis buffer directly to the cells or
tissues. After homogenization, the
lysates are transferred to DNA
columns. Thus the time-consuming
steps of material preparation are
eliminated.
The
DNA-bindingspecific matrix in the DNA column
allows the high efficient binding of
DNA, while RNA, proteins and other
cellular components are passed
through the DNA columns and are
collected. Genomic DNA are then
purified from the DNA columns,
while total RNA are purified from the
collected eluate through RNA
binding columns. Purified DNA can
be directly used for downstream
applications such as PCR, Southern
blotting, sequencing and restriction
enzyme digestion, while purified
total RNA can be used for RT-PCR,
Northern blotting or microarray
analysis. The working flow chart of
the kit is shown in Fig 1.
Cells or Tissue
Homogenize
and Lyse
Grind and
Lyse
Homogenize
with shredder
Kit content
Catalog No.
DR01-01
DR01-02
Storage
Preps
50
250
-
DNA columns
50
250
RT
RNA columns
50
250
RT
GD Lysis Buffer
60 ml
260 ml
RT
DNA Wash Buffer
12 ml
50 ml
RT
DNA Elution Buffer
10 ml
25 ml
RT
RNA Wash Buffer
12 ml
50 ml
RT
DEPC H2O
10 ml
25 ml
RT
100
500
RT
1
1
-
Collection tubes
Manual
Genomic DNA
Total RNA
Wash DNA
Add Ethanol
Caution! GD Lysis Buffer contains Chaotropic salts. Wear gloves when
handling this solution.
Specifications:
Elute DNA

Tissue and cells: up to 30 mg of tissues or 1-5x106 cells can be processed by using the kit

DNA sizes: up to 40 kb of DNA can be purified from this kit . The
purity of the DNA can be measured with expecting A260/A280≥1.8

DNA yield depends on the biological materials used. Up to 30 μg of
genomic DNA can be eluted
Bind RNA
Wash RNA
Buffer preparation
Elute RNA
Fig 1. DNA/RNA Miniprep kit flow chart
It is strongly advised that you familiarize yourself with the entire booklet
before starting.

Add β-mercaptoethanol to the Lysis Buffer to 1% (v/v) before use,
then store the buffer at 4oC.

Add 96-100% ethanol to DNA wash buffer bottle as follow:GD0101: Add 48 ml absolute ethanol (96-100%) to each bottle GD0102:Add 200 ml absolute ethanol (96-100%) to each bottle
Preheat the Elution Buffer to 70oC will increase the yield.

EasyPrepTM DNA/RNA Miniprep
3
4
EasyPrepTM DNA/RNA Miniprep
Protocol for Tissues
Storage and Stability
The EasyPrepTM DNA/RNA kit components are guaranteed for at least 12
months from the date of purchase.
Materials supplied by users:
 Homogenizer
 β-mercaptoethanol
 96-100% ethanol
 1.5 ml nuclease-free microcentrifuge tubes
 High speed microcentrifuge
I. Disruption and homogenization of tissue samples
It is critical to disrupt and homogenize the samples completely and
properly for high quality genomic DNA and RNA yield. The purpose for
homogenization is to reduce the viscosity by shearing genomic DNA and
other high molecular weight cell components to create a homogenous lysate. Incomplete homogenization may result in clogging the column and
reducing the RNA yield.
a. Sample disruption by mortar and pestle


Excise 10 mg tissues and freeze in liquid nitrogen immediate.
Grind the sample with ceramic mortar and pestle to a fine powder
under liquid nitrogen.
 Transfer the suspension into a tube pre-chilled in liquid nitrogen
and allow the liquid nitrogen to evaporate while the samples remain frozen.
 Add 350 μl GD Lysis Buffer before the sample gets thawed.
 Homogenize through a 20-gauge needle fitted to a syringe 5
times.
b. Rotor-Stator homogenizer for sample disruption and homogenization
Using a blade to slice tissue, weigh out 10 mg of animal tissue and
transfer it to a microcentrifuge tube, add 350 μl GD Lysis Buffer (with
β-mercaptoethanol added), homogenize the tissue according to the
manufacturer’s instruction. Using a proper size probes and generator,
the process simultaneously disrupts and homogenizes most of samples.
c.
Bead milling for sample disruption and homogenization
Cells and tissues can be disrupted and homogenized by rapid agitation in the presence of glass beads in 350 μl GD Lysis Buffer (with βmercaptoethanol added). Use 4-8 mm glass beads for animal tissues.
II. DNA purification
1. Centrifuge the homogenized tissue lysate at 11,000 rpm for 5
min. Transfer the supernatant to a gDNA binding column. Don’t
disturb the pellet during transfer. Centrifuge at 11,000 rpm for 30-60
sec. Keep the flowthrough for RNA purification. Place the DNA
column to a new collection tube.
2. Add 500 μl GD Lysis Buffer to the DNA column, centrifuge at
11,000 rpm for 30-60 sec. Discard the flowthrough.
EasyPrepTM DNA/RNA Miniprep
5
6
EasyPrepTM DNA/RNA Miniprep
3. Add 700 μl DNA Wash Buffer to the column, centrifuge at
11,000 rpm for 30-60 sec. Discard the flowthrough. Make sure
that ethanol is added to washing buffer as instructed before use.
4. Centrifuge additional 30-60 sec at 11,000 rpm. It is critical to
remove residual ethanol for optimal elution. Residual ethanol may
interfere subsequent applications.
Protocol for Cells
I.
DNA purification
1. Cell preparation and lysis:
5. Place the DNA column in a clean 1.5 ml microcentrifuge tube.
Add 50-100 µl pre-heated (70oC waterbath) DNA Elution Buffer
or Nuclease-Free water to the center of the column, incubate
for 1 min, centrifuge at 11,000 rpm for 30-60 sec to elute the
DNA. The purified DNA is stable in elution buffer or water if stored
at -20°C or below.
III. RNA purification

Adherent cells: scrape 1-5x106 cultured cells from the culture
dish or plate, pellet cells by centrifugation. Completely remove
the supernatant by aspiration.

Suspension cells: pellet 1-5x106 cells by centrifugation.
Completely remove the supernatant by aspiration.

Add 350 μl Lysis Buffer, vortex to resuspend the cell pellet.
Homogenize the lysate by passing it through a 20-gauge
needle fitted to a DNase free syringe for at least 5 times.
6. Add 350 μl 100% ethanol into the flow-through from Step 1; mix
thoroughly by pipetting up and down 10 times. Do not
centrifuge.
2. Transfer the homogenized lysate to a DNA column. Centrifuge
at 11,000 rpm for 30-60 sec. Keep the flowthrough for RNA
purification. Place the DNA column to a new collection tube.
7. Transfer the whole mixture to a RNA column and centrifuge at
11,000 rpm for 1 min. Discard the flow-through and put the
column back to the collection tube.
3. Add 500 μl Lysis Buffer to the column, centrifuge at 11,000 rpm
for 30-60 sec. Discard the flowthrough.
8. Add 500 µL RNA Prewash Buffer to the column and centrifuge
at 11,000 rpm for 30s. Discard the collection tube with the flowthrough. Put the column back to a new collection tube.
9. Add 500 µL RNA Wash Buffer to the column and centrifuge at
11,000 rpm for 30s. Discard the flow-through.
10. Centrifuge additional 30-60 sec at 11,000 rpm. It is critical to
remove residual ethanol for optimal elution. Residual ethanol may
interfere subsequent applications.
11. Place the RNA column in a RNase-free 1.5 ml microcentrifuge
tube. Add 35-50 µl DEPC-H2O to the center of the column,
incubate for 1 min, centrifuge at 11,000 rpm for 30-60 sec to
elute the RNA. The purified RNA is stable if stored at –70°C or
below.
4. Add 700 μl DNA Wash Buffer to the column, centrifuge at
11,000 rpm for 30-60 sec. Discard the flowthrough. Make sure
that ethanol is added to washing buffer as instructed before use.
5. Centrifuge additional 30-60 sec at 11,000 rpm. It is critical to
remove residual ethanol for optimal elution. Residual ethanol may
interfere subsequent applications.
6. Place the DNA column in a clean 1.5 ml microcentrifuge tube.
Add 50-100 µl pre-heated (70oC waterbath) DNA Elution Buffer
or Nuclease-Free water to the center of the column, incubate
for 1 min, centrifuge at 11,000 rpm for 30-60 sec to elute the
DNA. The purified DNA is stable in elution buffer or water if stored
at -20°C or below.
II. RNA purification
7. Add 350 μl 100% ethanol into the flow-through from Step 1; mix
thoroughly by pipetting up and down 10 times. Do not
centrifuge.
8. Transfer the whole mixture to a RNA column and centrifuge at
11,000 rpm for 1 min. Discard the flow-through and put the
column back to the collection tube.
9. Add 500 µL RNA Prewash Buffer to the column and centrifuge
at 11,000 rpm for 30s. Discard the flow-through.
EasyPrepTM DNA/RNA Miniprep
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8
EasyPrepTM DNA/RNA Miniprep
10. Add 500 µL RNA Wash Buffer to the column and centrifuge at
11,000 rpm for 30s. Discard the flow-through.
11. Centrifuge additional 30-60 sec at 11,000 rpm. It is critical to
remove residual ethanol for optimal elution. Residual ethanol may
interfere subsequent applications.
12. Place the RNA column in a RNase-free 1.5 ml microcentrifuge
tube. Add 35-50 µl DEPC-H2O to the center of the column,
incubate for 1 min, centrifuge at 11,000 rpm for 30-60 sec to
elute the RNA. The purified RNA is stable if stored at –70°C or
below.
Protocol for Whole blood, Serum and Plasma
The protocol is used to purify DNA and RNA from 100-200 μl whole
blood, serum or plasma. Whole blood in fresh, frozen or preserved
(such as in EDTA, heparin) forms can be used. If the samples can’t be
processed immediately, the samples can be preserved at 4 oC or –20oC
after addition of lysis buffer.
Trouble shooting
Problem
Possible reason
Suggested Improvement
Low A260/A280
ratios
Salt contamination
Add 2.5 volumes of ethanol and 0.1M
NaCl (final concentration) to precipitate
RNA. Incubate for 30 min at –20°C.
Centrifuge at 10,000 g for 15 min at 4°
Resuspend the RNA pellet in DEPCtreated water.
Low Yield
RNA in sample
degraded
Freeze samples immediately in liquid
nitrogen and store at -70°C after collect it.
Low Yield
The binding capacity of the membrane
in the spin column
was exceeded
Use of too much tissue sample exceeding the binding capacity of spin
column will cause the decreasing of
total RNA yield.
Low Yield
Ethanol not added
to buffer
Add ethanol to the RNA Wash Buffer
before purification.
The sample may
contain too much
genomic DNA.
Reduce the amount of starting tissue in
the preparation of the homogenate.
Most tissues will not show a genomic
DNA contamination problem at 30 mg
or less per prep. Reduce cell numbers
to 1x106 or increase buffer volume and
do multiple loadings to column.
Genomic DNA
contamination
1. Add 350 μl Lysis Buffer to the 100 μl whole blood, serum or
plasma, mix completely by vortexing 5 sec, incubate for 5 min.
Then follow the Step 2 and the rest of the protocol for cells.
Limited use and warranty
This product is warranted to perform as described in its labeling and in
Bioland’s literature when used in accordance with instructions. No other
warranties of any kind, express or implied, including, without limitation,
implied warranties of merchantability or fitness for a particular purpose,
are provided by Bioland. Bioland’s sole obligation and purchaser’s
exclusive remedy for breach of this warranty shall be, at the option of
Bioland, to replace the products, Bioland shall have no liability for any
direct, indirect, consequential, or incidental damage arising out of the
use, the results of use, or the inability to use it product.
EasyPrepTM DNA/RNA Miniprep
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10
EasyPrepTM DNA/RNA Miniprep
Other related products
Catalog #
Product Name
Preps
R01-01
RNA miniprep kit
50
R01-02
RNA miniprep kit
250
R02-01
Blood RNA miniprep kit
50
R02-02
Blood RNA mini kit
250
R03-01
Plant RNA mini kit
50
R03-02
Plant RNA mini kit
250
R96-01
96-well tissue RNA kit
4x96
R96-02
96-well tissue RNA kit
20x96
R01-03
RNA miniprep plus kit
50
R01-04
RNA miniprep plus kit
250
R01-05
RNA miniprep kit with DNase I
50
R01-06
RNA miniprep kit with DNase I
250
GD01-01
Genomic DNA miniprep kit
50
GD01-02
Genomic DNA miniprep kit
250
Bioland Scientific LLC
14925 Paramount Blvd., Suite C
Paramount, CA 90723
USA
Tel:
(877) 603-8882
Fax:
(562) 733-6008
Email: [email protected]
[email protected]
Visit our web at www.bioland-sci.com and learn more about
Bioland products
EasyPrepTM DNA/RNA Miniprep
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