DNA STAT -60
Key Benefits
*Isolates high quality genomic DNA in one hour from
tissues and cells with a single monophasic reagent
*Can be used to back extract DNA from "Single Step Method " RNA
Preps (i.e, RNAzol, RNA Bee, RNA Stat-60)
*Does not contain phenol
*RNA and proteins sequestered in organic and interphase; DNA remains
in upper aqueous phase.
Pricing
|
Catalog # |
Size |
Price |
|
TL-4200 |
50 ml |
$76.00 |
|
TL-4210 |
100 ml |
$108.00 |
|
TL-4220 |
200 ml |
$195.00 |
1. INTRODUCTION
DNA STAT-60, a single monophasic reagent containing a chaotropic
cell disrupter and a non-corrosive phenol free extraction reagent,
replaces cumbersome, labor intensive methods of genomic DNA
isolation. Following tissue or cell homogenization in the DNA
STAT-60, and after the addition of chloroform, the homogenate
separates into two phases: the aqueous phase and organic phase. The
DNA remains in the aqueous phase while RNA and other cellular
components, including proteins, are preferentially partitioned in
the organic phase and interface. The DNA STAT-60 method does not
require ultra centrifugation, and can be completed in under 1 hour.
The DNA STAT-60 isolates high molecular weight genomic DNA from
samples of human, animal, plant, yeast, and bacterial origin and is
particularly well suited for the simultaneous processing of multiple
samples. RNA and proteins are sequestered in organic and interface:
DNA remains in upper aqueous phase. Protocol can be completed in 60
minutes. Single monophasic reagent with extended shelf life. Does
not contain phenol or hazardous organics. Isolates genomic DNA from
samples of human. animal, plant, yeast and bacterial origin. Can be
used to back extract DNA from "SINGLE STEP METHOD" RNA
preps (i.e. RNAzol B. RNA STAT-60).
2.
REAGENTS SUPPLIED DNA STAT-60TM
50
ml, 100 ml, or 200 ml bottle containing clear solution of DNA
STAT-60
TM.
PREPARATION Ready to use.
STORAGE Refrigerate at 2-8oC. Protect from
exposure to light.
STABILITY 6 months. Refer to expiration date stamped on
label.
3. REAGENTS REQUIRED, BUT NOT SUPPLIED
Chloroform (ACS grade) Isopropanol (ACS grade) Ethanol (ACS
grade)
4. PROTOCOL
DNA isolation by the DNA STAT-60TM
method includes the following steps:
1. Homogenization DNA STAT-60 (1 ml per 50-100 mg tissue, or
5-10 x 106 cells).
2. DNA Extraction 1 vol. of homogenate + 0.2 vol. of
chloroform.
3. DNA Precipitation 0.5 vol. of isopropanol.
4. DNA Wash 75% ethanol. Unless stated otherwise the
procedure is carried out at room temperature.
4.1 HOMOGENIZATION
A. TISSUES: Homogenate tissues in the DNA STAT-60 (1 ml/50-100
mg tissue) in a glass-Teflon or polytron homogenizer.
B. CELLS: Cells grown in mono layer are lysed directly in a
culture dish by adding DNA STAT-60 (1 ml/3.5 cm petri dish) and
passing cell lysate 5-10 times through a pipette. Cells grown in
suspension are sedimented then lysed in a DNA STAT-60 (1 ml per 5-10
x 106 cells by repetitive pipetting.
4.2 DNA EXTRACTION Following homogenization,
add 0.2 ml of chloroform per 1 ml of DNA STAT-60, cover the sample
tightly, shake vigorously for 15 seconds and let it stay at room
temperature for 2-3 minutes. Centrifuge the homogenate at 12,000 g
(max.) for 15 minutes at 4oC. Following centrifugation
the homogenate separates into two phases: a lower organic phase and
the upper aqueous phase. DNA remains in the aqueous phase whereas
RNA and proteins are in the interface and the organic phase.
4.3 DNA PRECIPITATION
Remove supernatant and wash the DNA pellet once with 75%
ethanol by vortexing and subsequent centrifugation at 7,500 g (max.)
for 5 minutes at 4oC. Add at least 1 ml of 75% ethanol
per 1 ml of the DNA STAT-60 used for the initial homogenization. At
the end of the procedure, dry the DNA pellet briefly by air drying
(5-10 min.). Dissolve the DNA pellet in water or in 1 mM EDTA, pH 7.
Pass the pellet a few times through a pipette tip. An incubation for
10-15 minutes at 55-60oC may be required to dissolve DNA
samples.
5. RECOVERING DNA FROM "SINGLE STEP METHOD"
(RNAzol B, RNA STAT-60 PREPS).
5.1 DNA REVERSE EXTRACTION
Remove aqueous layer containing RNA. Add 800 ul of DNA STAT-60
reagent per 1 ml of RNAzol B or RNA STAT-60 used for the initial
homogenization. Add 0.2 ml of chloroform per 1 ml of DNA STAT-60,
shake vigorously for 15 seconds and let it stay at room temperature
for 2-3 minutes. Centrifuge the homogenate at 1000 g (min.)-12,000 g
(max.) for 15 minutes at 4oC. Following centrifugation
the homogenate separates into two phases: a lower organic phase and
the upper aqueous phase. DNA remains in the aqueous phase whereas
RNA and proteins are in the interface and organic phase.
5.2 DNA PRECIPITATION
Follow procedure as outlined in section 4.4.
6. SPECIAL HANDLING PRECAUTIONS
The DNA STAT-60 contains an irritant (Guanidinium Salts). Can be
fatal. When working with DNA STAT-60 use gloves and eye protection
(shield, safety goggles). Do not get on skin or clothing. Avoid
breathing vapor. Read warning note on bottle.
REFERENCES
1. Chomczynski, P. and Sacchi, 1987. Single Step Method and RNA
Isolation by Acid Guanidinium Thiocyanate-Phenol-Chloroform
Extraction. Biotechniques 8:148-149.
2. Maniatis, T., E.T. Fritsch and J. Sambrook, 1982. Molecular
Cloning: A Laboratory Manual.
3. Winberg. G., 1991. A Rapid Method for Preparing DNA from Blood.
Suited for PCR Screening of Transgenes in Mice. PCR Methods Applic.
1:72-74. |
+32 1658 9045
+32 1650 9045
