Binding liquid and superhelix plasmid large extraction kit based on binding liquid

Abstract

The invention discloses a binding solution and a superhelix plasmid large extraction kit based on the binding solution. The binding solution comprises 1-5 M of guanidine isothiocyanate, 10-30% (V / V) polyethylene glycol 8000, 60-90 mM of Tris-HCl, 20-40 mM of EDTA and 0.5-2 M of NaCl, and the PH is 4-7. According to the superhelical plasmid large extraction kit based on the binding liquid, firstly, escherichia coli containing target plasmids is subjected to resuspension, splitting decomposition and neutralization through an alkali splitting decomposition method; adding a binding solution, uniformly mixing, and standing at room temperature to form a mixed solution; centrifuging, filtering the mixed solution through a centrifugal column, and discarding the filtrate; and finally, washing and eluting the plasmids adsorbed on the centrifugal column to obtain the target plasmids. The special binding liquid can enhance the binding efficiency of the plasmids and the centrifugal column and improve the content of the extracted superhelical plasmids.

Description

technical field [0001] The invention relates to the technical field of plasmid extraction, in particular to a supercoiled plasmid large-scale extraction kit. Background technique [0002] Plasmid is a genetic unit capable of autonomous replication outside the chromosome, and bacterial plasmid is one of the most commonly used vectors for recombinant DNA technology. The self-replicating ability of plasmids and the genetic information they carry make them very practical and valuable in genetic engineering research and its downstream applications. Plasmid extraction is one of the most critical steps in genetic engineering technology, which is related to the smooth progress of subsequent transformation, DNA sequencing, PCR, enzyme digestion and other experiments. [0003] At present, the improved alkaline lysis method is the most commonly used and most effective plasmid extraction method. The basic principle is that when the bacteria are lysed in NaOH and SDS, the protein and DN...

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Problems solved by technology

[0005] At present, the large plasmid extraction kits or reagents developed by some reagent companies at home and abroad have pollution problems: first, the extracted plasmid bacterial genomic DNA is contaminated, which will have adverse effects on downstream experiments; second, it needs to be added during the extraction process. Toxic reagents such as chloroform-isopropanol have potential safety hazards; the third is that there are few supercoiled plasmids extracted

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