Manual

Product Number: PLK2001

Shipping and Storage

1. When using for the first time, add all RNaseA carried by the reagent kit to Buffer P1 (final concentration 100μg/ml) and store at 4℃. If RNaseA is inactivated in Buffer P1, the extracted plasmid may contain trace amounts of RNA residue. Adding RNase A to Buffer P1 is sufficient.
2. Buffer ER can be stored at 4℃ for one month. If it needs to be stored for a long time, it is recommended to store it at -20℃!
3. When the ambient temperature is low, SDS in Buffer P2 may precipitate and become turbid or precipitate. It can be heated in a 37℃ water bath for a few minutes to restore clarity. Do not shake violently to avoid excessive foam formation.
4. To avoid volatilization, oxidation, and pH changes caused by prolonged exposure of reagents to the air, each solution should be covered tightly in a timely manner after use.

Components

Component	Storage	PLK2001 20Preps
RNaseA(10mg/ml )	-20℃	1.3ml
Buffer P1	4℃	130m1
Buffer P2	RT	100 ml
Buffer P3	RT	110 ml
Buffer IRA	RT	3 ml
Buffer IRB	RT	30 ml
Buffer ER	-20℃	10 ml

Description

The conventional centrifugal column plasmid extraction kit is not suitable for the extraction of large plasmid DNA such as BAC/PAC/P1/Cosmid. This is mainly because the molecular weight of large plasmid DNA is very large, often exceeding 100kb, and generally has low copy number and low yield. The use of conventional centrifugal columns to adsorb membranes has low adsorption efficiency and yield, and passing the membrane will interrupt these large molecular weight plasmid DNA. This kit uses an improved alkaline lysis method to extract plasmid DNA from cultured bacteria. With a unique solution formula and Buffer ER, it only requires a few simple centrifugations to remove impurities such as proteins, polysaccharides, endotoxins, RNA, and obtain high-quality plasmid DNA. The OD260/280 of purified DNA is usually around 1.8, and the resulting plasmid can be directly applied in tasks that require high DNA purity, such as cell transfection and even animal in vivo experiments. The purification process in the later stage is operated in a 1.5ml centrifuge tube, which is simple, does not require special equipment, does not require column passing, and does not require phenol chloroform extraction; Plasmids released by bacterial lysis can be completely recovered without worrying about the loss of plasmid DNA. This method extracts and purifies plasmid DNA with minimal damage to plasmids. Even large plasmids or ultra large BAC/PAC plasmids with a size of 100kb or even 200kb can be effectively purified as long as they can be extracted by alkaline lysis. In addition, the solution type reagents can be scaled up or down in proportion for small/medium/large extraction. Finally, any small volume can be chosen to dissolve the plasmid, with a concentration of up to 3μg/μl.

Features

1. No need to use toxic reagents such as phenol and chloroform, and no need for ethanol precipitation. Rapid and convenient extraction of high-purity BAC plasmid DNA with a typical yield of 30-50μg can be achieved from 150ml of Escherichia coli LB (Luria Bertani) culture medium, with an extraction rate of 80-90%.
2. The obtained plasmids have high yield, high superhelix ratio, and good purity, and can be directly used for various molecular biology experiments such as transfection, sequencing, and library.
3. The endotoxin content is extremely low (<0.1EU/μg DNA) and can be directly applied to cell transfection

Application

Suitable for the preparation of large plasmids such as BAC/PAC/P1/Cosmid

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