iNtRON Biotechnology

Plasmid & Probe DNA

Genomic DNA Prep

RNA Prep

Protein

PCR

RT/RT-PCR

DNA & Protein Markers

Chemical & Other

Cloning & Enzyme

Molecular Diagnostics

HOME > Plasmid & Probe DNA > Plasmid DNA > DNA-midi™ SV Plasmid DNA Purification Kit

070410_easy_llp_1

DNA-midi™ SV Plasmid DNA Purification Kit provides easy and rapid method for the midi scale preparation of plasmid DNA from bacterial cells. This kit can be used to isolate and purify any plasmid, also can isolated maximum 40 kb size plasmid DNA. The plasmid DNA is free from protein, genomic DNA, and RNA contaminants. This pure plasmid DNA is ready for PCR, cloning, automated or manual sequencing, transfection, synthesis of labeled hybridization probes, electroporation, and enzymatic restriction analysis.

Easy to use - organic extraction or ethanol precipitation is no required.
No phenol or chloroform is used.
Spend only 30 min (vacuum protocol), 50 ~ 60 min (spin protocol) to extract plasmid DNA
Cell lysates remove easily with Pre Column.
:After mixing with M3 Buffer, the cellular debris and precipitates should be removed completely not to clog Binding Column in subsequent binding. Pre Column facilitates the clearance of the lysate by filtration instead of laborious incubation on ice and centrifugation which has been used widely in traditional methods.
Plasmid DNA binds selectively to silica membrane.
This column system applies spin and vacuum protocol.

- Store at RT except for M1 Buffer. M1 Buffer should be stored at 4℃ after adding RNase A Solution.

M1 Buffer (Resuspension Buffer) : Before use, add 4 ㎖ of RNase A solution to M1 Buffer. Then, store at 4℃.	60 ml
M2 Buffer (Lysis Buffer) : Check M2 Buffer for SDS precipitation due to low storage temperature, in which case it is necessary to dissolve the SDS by warming to 37℃.	60 ml
M3 Buffer (Neutralization Buffer)	60 ml
Washing Buffer A : endA⁺ strains such as HB101, the JM series strains, PR series strains and some other wide-type stains have high endonucleases activity. Endonucleases that can degrade plasmid DNA are essentially removed by Washing Buffer A of DNA-midi^TM SV Kit.	300 ml (150 ml x 2ea )
Washing Buffer B : Before use, if 40 ㎖ of Washing Buffer B's bottle, add 160 ㎖ of absolute EtOH. And if 20 ㎖ of Washing Buffer B's bottle, add 80 ㎖ of absolute EtOH.	60 ml (40 ml x 1ea / 20 ml x 1ea)
Elution Buffer : DNase / RNase free Ultra-Pure solution.	30 ml
RNase A Solution (10mg/ml) : After receiving, store at 4℃.	4 ml
Pre Column (Clear Color) : Inserted into the 50 ㎖ disposable tube (collection tube)	25 Col.
Binding Column (Yellow Color) :Inserted into the 50 ㎖ disposable tube(collection tube)	25 Col.

Shape and application of Pre & Binding Columns

Fig. 1. Shape and application of Pre & Binding Column
(a) Normal shape of Pre-filtration column & Binding column
Yellow color : Binding Column, Clear color : Pre Column
(b) Shape of 50 ㎖ tube application (Protocol A)
(c) Shape of vacuum manifold application (Protocol B)

Tendency of plasmid DNA yield & purity from plasmid size

Fig. 2. Gel analysis of various size plasmid DNA
Lane M, 1kb ladder;
Lane 1, pUC18 (2.9 kb);
Lane 2, pTA (7.1 kb); L
Lane 3, pCEP4 (10.5 kb);
L ane 4, pAdEASY-1 (33.2 kb);
Lane 5, pJM17 (40.2 kb)

[ Table. 1 ] Yield & purity of various size plasmid DNA isolated from DH5α

Plasmid DNA isolated from various E.coli

Fig. 3. Gel analysis of plasmid DNA isolated from various E. coli host strain
Lane M, 1kb Ladder DNA Marker; lane 1,DH5a; lane 2, JM109; lane 3, TOP10; lane
4, BL21; lane 5, BL21(DE3) ; lane 6,BL21(DE3)pLysS

[Table. 2] Yield & purity of plasmid DNA isolated from various E.colihost strain

Transfection & Luciferase assay

Fig 4. Transfection & Luciferase assay in K-562 and 293T

타사 비교 DATA

Fig.6. 타사 비교 DATA
Lane M, 1 kb ladder;
L ane 1, iNtRON; Lane 2, Supplier A;
Lane 3, Supplier B; Lane 4, Supplier C

[ Table. 3] Yield & purity of plasmid from iNtRON and other companies kit

Elution volume에 따른 plasmid DNA yield & purity

Fig. 7. Gel analysis of plasmid DNA from various elution volumne.
Lane M, 1 kb Ladder DNA Marker;
Lane 1,250ml; Lane 2, 500 ml; Lane 3, 750 ml;
Lane 4, 1 ml; Lane 5, 1.5 ml; Lane 6, 2 ml

Fig. 8. The overall yield and concentraion of plasmid DNA depending on the elution volumen.