Description : Key to the kit is our proprietary DNA binding systems that allow the high efficient binding of DNA to our ezBindTM matrix while proteins and other impurities are removed by Wash Buffer. Nucleic acids are easily eluted with sterile water or Elution Buffer. Unlike other kits in the markets, our patented plasmid purification kit has no chaotropic salts in the buffer, the purified DNA is guanidine/anion exchange resin residues free which enable the high performance of downstream applications such as transfection, restriction mapping, library screening, sequencing, as well as gene therapy and genetic vaccinations. Copy numbers: The yield of plasmid DNA depends on the origin of the replication and the size of the plasmid. The protocols are optimized for high copy number plasmid purification. For low copy number plasmids, both the culture volume and the buffer volume need to be scaled up 2 to 3 times. Reference the table below for the commonly used plasmids, Plasmid Origin Copy Numbers Expected Yield(µg per 200 mL) pSC101 pSC101 5 12 pACYC P15A 10-12 25-40 pSuperCos pMB1 10-20 30-50 pBR322 pMB1 15-20 35-50 pGEMR Muted pMB1 300-400 350-450 pBluescriptR ColE1 300-500 450-600 pUC Muted pMB1 500-700 700-1,000 Host Strains: The strains used for propagating plasmid have significant influence on yield. Host strains such as JM101, JM110, HB101, TG1 and their derivatives, normally have low plasmid yield due to either endogenous endonucleases or high carbohydrates released during lysis. We recommend transform plasmid to an endA- strain if the yield is not satisfactory. For purifying plasmid DNA from endA- strain such as Top 10, DH5a, and C600, we recommend use product number PD1511. Optimal Cell Mass (OD600 x mL of Culture): This procedure is designed for isolating plasmid grown in standard LB medium (Luria Bertani) for 12 -16 hours to a density of OD600 2.0 to 3.0. If rich medium such as TB or 2xYT are used, make sure the cell density doesn't exceed 3.0 (OD600). A high ratio of biomass over lysis buffers result in low DNA yield and purity. The maxi column has an optimal biomass of 450-550. For example, if the OD600 is 2.5, the optimal culture volume should be 200 mL. Culture Volume: Use a flask or tube with a volume at 4 times the culture medium to secure optimal condition for bacteria growth. Don't exceed the maximum culture volume suggested in the protocol. Incomplete lysis due to over amount of bacterial culture results in lower yield and less purity. Buffer A1 should be stored at 4℃ once RNase A is added. All other materials can be stored at room temperature. The Guaranteed shelf life is 18 months from the date of purchase. Prepare all components and get all necessary materials ready by examining this instruction booklet and become familiar with each step. Catalog # PD1713-01 PD1713-02 Preps 10 25 ezBindTM Columns 10 25 Buffer A1 110 mL 270 mL Buffer B1 110 mL 270 mL Buffer C1 135 mL 330 mL EndoClean Buffer 5 mL(Optional: For Endofree) 10 mL(Optional: For Endofree) Buffer KB 120 mL 270 mL DNA Wash Buffer 36 mL 80 mL RNase A 11 mg 27 mg Elution Buffer 30 mL 60 mL EndoFree Water 4 mL(Optional: For Endofree) 10 mL(Optional: For Endofree)Introduction
Important Notes
Storage and Stability
Before Starting
Important
Materials supplied by users
Kit Contents
- Model: PD1713-01
- Manufactured by: Biomiga