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High efficiency plasmid extraction kit
Plasmid Miniprep Kit is an innovative system that radically simplifies extraction and purification of nucleic acids from a variety of sources. The Plasmid Miniprep Kit combines the power of silica membrane technology with the time-tested consistency of alkaline-SDS lysis of bacterial cells to deliver high quality DNA in under half an hour. The silica membrane reversibly, binds DNA or RNA under certain optimal conditions allowing proteins and other contaminants to be removed. Nucleic acids are easily eluted with deionized water or low salt buffer. Plasmid Miniprep Kit mini-columns facilitate the binding, washing, and elution steps thus enabling multiple samples to be simultaneously processed. Yields vary according to plasmid copy number, E.coli strain, and conditions of growth, but 14.5ml of overnight culture in LB medium typically produces 140µg plasmid DNA. The product is suitable for automated fluorescent DNA sequencing, restriction endonuclease digestion, and other manipulations.
Kit Content, Storage and Stability

GS1011 (100 preps)
GS1012 (200 preps)
RNase A (10mg/ml)
Buffer P1
25 ml
50 ml
Buffer P2
25 ml
50 ml
Buffer P3
35 ml
70 ml
Buffer PE
31.5 ml
63 ml
Add 18.5ml absolute ethanol before first use
Add 37ml absolute ethanol before first use
Buffer WB
25 ml
50 ml
Add 100ml absolute ethanol before first use
Add 200ml absolute ethanol  before first use
Buffer EB
15 ml
20 ml
DNA Spin Column
100 pcs
200 pcs
Collection Tube2ml
100 pcs
200 pcs

All reagents are stable for 24 months at RT, when stored properly.
1. If RNaseA is inactive, RNA will contaminate the plasmid. Add additional RNaseA to Buffer P1.
2. Check Buffer P2 for SDS precipitation due to low storage temperatures. If necessary, dissolve the SDS by warming to 37C
3. Please ensure the bottles of buffer tightly capped when not in use to prevent reagents evaporating, oxidation and pH changing.
4. Dilute Buffer WB with three volume absolute ethanol before start.
(1)Rapid and convenient. Do not contain poisonous phenol etc and not need carrying ethanol precipitation. Multi-elution can ensure high-purified DNA, which can be applied to all kinds of molecular experiments such as PCRSouthern-blot and digestions directly.
(2)The silica membranes in the spin-column come from the world-famous company.
(3)Unique content can effectively remove the nuclease, even apply to rich-nuclease stains of JM and HB101,
Please read this section before your experiment.
1. All the centrifugation can be performed at room temperature.
2. Buffer P3 includes the stimulating compound. Please wear latex gloves, avoiding skin, eyes and cloth to be contaminated. If that, please use water or physiological saline washing.
3. The yield of plasmid is related with concentration of liquid culture and copy number. For high copy plasmids, picking a single colony from a freshly streaked selective plate, inoculating in 1.5-4.5 ml LB medium containing the appropriate selective antibiotic, shaking over night at 37C, the yield of plasmid may achieve 20g.For low copy plasmids or size>10kb plasmids, we recommend collecting 5-10ml overnight culture and scaling up volumes of buffer P1, P2 and P3.
4. DNA concentration and quality can be determined by UV and agarose gel electrophoresis. 1OD260 may be 50g/ml DNA. Typically, the majority of the eluted DNA is in monomeric supercoil form, sometimes they may display different types, single or two even more bands in agarose gel electrophoresis because of influenced by culture time and operations of extracting.
5. If want to know the size of the plasmid, please make the restriction endonuclease digestion to get the exact size compared with the DNA Marker.
6. No EDTA in Buffer EB, which will have no influence on down-stream reactions. Also you can use water when eluting, but please ensure pH>7.5 and store at -20. If for long-term storage, dissolve plasmid in TE10mM Tris-HCl 1mM EDTApH 8.0. Because EDTA will affect the down-stream reactions, dilute the solution before use.
Before Starting
Add the all the provided RNase A to Buffer P1 before use, to give a final concentration of 100g/ml. Store the P1/RNase A mixture at 4.
Dilute BufferWB and Buffer PE with absolute ethanol, vortex adequately, then mark the check box, avoid multi-adding!
1.Harvest 1.5-4.5ml overnight culture fluid, centrifugation at 9,000rpm for 30s. Collect bacterial pellet, discard the supernatant.
2.Resuspend the bacterial pellet by adding 250l Buffer P1, and vortex. Complete resuspension (no visible cell clumps) of cell pellet is vital for obtaining good yields.
3.Add 250l Buffer P2 and gently mix by inverting and rotating tube 6-10 times to obtain a clear lysate. Avoid vigorous mixing as this will resulting in shearing chromosomal DNA and lower plasmid purity. Do not allow the lysis reaction to proceed more than 5 min as this will damage plasmid.
4.Add 350l Buffer P3 and immediately mix by inverting and rotating tube 6-10 times. Incubate at room temperature for 5min. Centrifuge at 13,000rpm for 10min at room temperature.
5.Add the clear supernatant carefully into Spin-column AC. Centrifuge at 13,000 for 1min. Discard the flow-through liquid.
6.Add 500l Buffer PE. Centrifuge for 30-60s at 13,000rpm. Discard the flow-through.
7.Add 700l Buffer WB. Centrifuge for 30-60s at 12,000rpm. Discard the flow-through.
Note: Buffer WB must be diluted with absolute ethanol before first use.
8.Add 500l Buffer WB. Centrifuge for 30-60s at 12,000rpm. Discard the flow-through.
9.Centrifuge the empty column at 13,000rpm for 2 min. Air-dry for 3-5 min at room temperature.
10.Transfer the Spin-column AC to a clean1.5ml microcentrifuge tube, add 60-100l Buffer EB (water bath in 65-70 before use) directly onto the silica-membrane. Incubate 1 min at room temperature. Centrifuge at 13,000rpm at 1 min.
The volume of elution buffer could be adjusted according to needs. Appropriately reduce elution volume can increase concentration. But the minimum volume is 50l, too less will decrease the elution efficiency and the DNA yield.
11.Keep DNA at 2-8 (-20 for long-term storage).
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