[Product Introduction]
Magnetic bead method plasmid DNA small amount extraction kit is a bacterial plasmid DNA small amount extraction kit recently developed by Suzhou Beike Zhenze Biotechnology Co., LTD. This product combines magnetic nano separation technology with SDS alkali lysis method to extract high quality plasmid DNA from bacterial cells. The cell fragments and SDS complex precipitate under centrifugal force. Under certain conditions, the magnetic beads can adsorb plasmid DNA in the supernatant well, and other impurities such as proteins and salt ions can be removed by washing. When conditions change, the beads release the adsorbed plasmid DNA. For high-copy plasmids, more than 15 μg plasmid DNA was obtained from 2 mL of overnight culture (OD600 = 2.0). The plasmids extracted by this kit are of high purity, and the OD260/OD280 ratio is generally between 1.7 and 1.9, and the OD260/OD230 ratio is greater than 2.0, which can be directly used for subsequent experiments such as transformation, DNA sequencing, PCR, PCAR-based mutation, in vitro transcription and enzyme digestion.
【 characteristics 】
L For high-copy plasmids, over 15 μg plasmid DNA was obtained from 2 mL of overnight culture (OD600 = 2.0);
L Compared with similar products, the extraction efficiency is higher, the purity of the obtained plasmid is higher;
L Compared with the column method, the centrifugation times can be greatly reduced, and the plasmid with better integrity can be obtained.
Multiple samples can be processed simultaneously to realize high-pass quantification and automation of plasmid extraction.
[Preservation methods and Precautions]
Please store the Beads and RNase A in the kit at 2-8℃ and the rest of the reagents at room temperature. See the packing for the expiry date.
L Freezing and centrifugation are strictly prohibited. Freezing and centrifugation may cause irreversible damage to magnetic beads.
L Magnetic beads will gather into clusters after long-term placement, thus reducing the surface area of magnetic beads and reducing the recovery rate of samples. It is necessary to thoroughly mix magnetic beads by eddy oscillation before use (usually 20 seconds of eddy oscillation).
[Kit composition]
component
100 times
save
Resuspension solution
20 ml
The normal temperature
Lysis buffer
20 ml
The normal temperature
Neutralization solution
20 ml
The normal temperature
Wash buffer
75% ethanol (for users)
The normal temperature
Elution buffer
10 ml
The normal temperature
bead
2 ml
2-8 ℃
RNase A
2 ml
2-8 ℃
Isopropyl alcohol
Users should bring along their own
The normal temperature
DNA concentration and purity detection :(1) the size of the obtained genomic DNA fragments was related to factors such as sample preservation time and shear force during operation. The concentration and purity of the obtained DNA fragments could be detected by agarose gel electrophoresis and ultraviolet spectrophotometer. (2) DNA should have a significant absorption peak at OD260, and OD260=1 is equivalent to about 50ng/ul double-stranded DNA and 40ng/ul single-stranded DNA. The OD260/OD280 ratio should be 1.7~1.9. If deionized water is used in elution, the ratio will be low, because pH and ions will affect the absorbance, which does not mean low purity.
[Operation Procedure]
Points bacteria -
1) Take 1~ 1.5ml of escherichia coli cultured overnight and put it in a 2 mL microcentrifuge tube, centrifuge it at 4℃ for 0.5 minutes at 9000 RPM, and blot out the culture medium as much as possible;
Suspended bacteria -
2) add 200 ul Resuspension solution and 20ul RNaseA suspended bacteria to mix well;
The cracking -
3) Add 200 UL Lysis buffer, close the mouth of the tube, gently invert the centrifuge tube 4-6 times to make the solution transparent and thick, and let stand at room temperature for 3-5min until complete Lysis.
Neutralizing -
4) Add 200 ul ice-precooled Neutralization solution, cover the centrifugal nozzle tightly and gently reverse 4-6 times, and add 50 ul isopropyl alcohol.
In combination with -
5) Centrifugation at 13000 RPM for 5 min at 4 ℃, carefully absorbing 500ul supernatant to the new 1.5 mL centrifuge tube (300 ul isopropanol and 20 ul PuriMag Bead were pre-mixed in the centrifuge tube); Blow 3~5 times, set for 5 min; Place the centrifuge tube on a magnetic rack for 10 seconds and carefully discard the supernatant.
The washing -
6) Add 700 UL Wash buffer into the centrifuge tube, blow and mix with pipette gun, put the centrifuge tube on a magnetic rack for 10 s, and carefully discard the supernatant. Repeat this step once. The centrifuge tube was inverted for 2~3 min, and then dried at room temperature for 5 min to allow the residual ethanol to volatilize clean, so as not to affect subsequent experiments.
Des -
7) Add 50-100 ul Elution buffer into the centrifugal tube, blow and mix with pipette gun for 1-2 min. (Elution buffer in 65-70 ℃ after preheating Elution effect is better, reduce the Elution volume or multiple Elution can increase the DNA concentration) Put the centrifugal tube on the magnetic rack for 10 s, carefully suction up the natant into the new centrifugal tube, that is, the extracted plasmid DNA, can be stored at 2~8 ℃, Long-term storage should be placed at -20 ℃.
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Warm tip: the products supplied by Beijing Beike Xincai Technology Co., Ltd. are only used for scientific research, not for human body |
| Item ID |
CAS |
ID |
Pack |
Parameter |
Stock |
Make up |
Price |
| BK2021081231 |
|
BK2021081231 |
|
|
100 |
|
$0 |
