Endotoxin Free Midi Kit (PIFE25)

Midiprep Plasmid Kit Endotoxin Free
Endotoxin Free Midi Kit (PIFE25)

The Presto™ Midi Plasmid Kit Endotoxin Free was designed for Plasmid DNA Purification and endotoxin removal from 50-150 ml of cultured bacterial cells using an efficient plasmid midiprep system. For processing smaller volumes, the Presto™ Mini Plasmid Kit is also available. Both plasmid kits include TrueBlue Lysis Buffer (an optional color indicator) to prevent common handling errors, while ensuring efficient cell lysis and neutralization. The Presto™ Midi Plasmid Kit Endotoxin Free uses a modified alkaline lysis method and RNase treatment  to obtain clear cell lysate with minimal genomic DNA/RNA contaminants. Using an efficient gravity-flow procedure, plasmid DNA in the crude lysate is bound by the plasmid midi column and the contaminants are removed with PW Buffer. The purified plasmid DNA is eluted by a high salt buffer and then precipitated with Isopropanol for desalting. The entire procedure can be completed in 80 minutes without ultracentrifuges, HPLC or other toxic reagents.  

Advantages (Cat. # PIFE25)
  • High Yield: >400 µg of pure transfection grade plasmid DNA from 100 ml of cultured bacterial cells
  • Efficient endotoxin removal (<0.1 EU/µg DNA) verified by LAL
  • Convenient: TrueBlue Lysis Buffer is an optional color indicator included with this plasmid kit in order to reduce common handling errors, while ensuring efficient cell lysis and neutralization and maximum plasmid product yield so you can feel confident with each and every plasmid prep.
  • Sample Volume: 50-150 ml of cultured bacterial cells
  • Format: Plasmid Midi Anion-Exchange Resin Columns and Filter Columns (membranes optimized for plasmid DNA purification)
  • Elution Volume: 500 µl - 2 ml
  • Storage: dry at room temperature (15-25ºC)
  • Operation time: within 80 minutes (only 40 minutes when combined with the Presto™ Plasmid Concentrator)
Applications
 
Transfection, Sequencing, Ligation, PCR, In-vitro Transcription, Microinjection, Restriction Enzyme Digestion, Gene Gun
 
Components
  • PM1, PM2, PM3 Buffers
  • PER Buffer (Endotoxin Removal Buffer)
  • TrueBlue Lysis Buffer
  • PEQ Buffer
  • PW Buffer
  • PEL Buffer
  • RNase A
  • Filter Columns (yellow spin columns)
  • Plasmid Midi Columns (anion exchange white resin columns)
Quality Control
 
The quality of the Presto™ Midi Plasmid Kit Endotoxin Free is tested on a lot-to-lot basis according to Geneaid's ISO-certified quality management system. Plasmid DNA from a 50 ml overnight E. coli (DH5α) culture, containing plasmid pBluescript (A600 > 2 U/ml) is purified using the Presto™ Midi Plasmid Kit. Following the purification process, a yield of more than 200 μg is obtained and the A260/A280 ratio is between 1.8-2.0. The purified plasmid (1 µg) is used in EcoRI digestion, and analyzed by electrophoresis.
 
Presto™ Midi Plasmid Kit Endotoxin Free Functional Test Data
Brand
Test
260/280
260/230
Yield
Average
Geneaid
(Lane 1, 2)
50 ml
1.85
2.25
273.00 µg
265.90 µg 
50 ml
1.87
2.23
258.80 µg
100 ml
1.87
2.30
373.20 µg
391.20 µg
100 ml
1.87
2.14
409.20 µg
Competitor
(Lane 3, 4)
50 ml
1.85
2.26
121.10 µg
131.30 µg
50 ml
1.85
2.29
141.50 µg
100 ml
1.86
2.32
285.80 µg
287.85 µg
100 ml
1.87
2.33
289.90 µg
Figure 1. Plasmid DNA was extracted using both the Presto™ Midi Plasmid Kit Endotoxin Free (lane 1,2) and the equivalent competitor's plasmid midi kit (lane 3,4). The purified supercoiled plasmid DNA [50 ml and 100 ml overnight E. coli (DH5α) culture, containing a 3 kb plasmid pBluescript (A600 > 2 U/ml, OD600 = 3.8)], was used in EcoRI digestion and analyzed by electrophoresis on a 1% agarose gel.
M = Geneaid 1 Kb DNA Ladder, Lane 1: Presto™ Midi Plasmid Kit Endotoxin Free (50 ml), Lane 2: Presto™ Midi Plasmid Kit Endotoxin Free (100 ml), Lane 3: Equivalent Competitor Kit (50 ml), Lane 4: Equivalent Competitor Kit (100 ml)
Column Types Included With the Presto™ Midi Plasmid Kit Endotoxin Free
Filter Column              
Figure 2. This centrifuge midi plasmid spin column is used to filter the neutralized colorless mixture and remove contaminants to facilitate DNA binding to the white membrane of the Plasmid Midi Column.
Plasmid Midi Column
Figure 3. This gravity flow plasmid midi anion exchange resin column is used for binding, washing and eluting plasmid DNA.   
TrueBlue Lysis Buffer Optional Color Indicator
Cell Lysis
Neutralization
Figure 4. TrueBlue Lysis Buffer is an optional color indicator included with this Plasmid Kit to eliminate common handling errors and ensure efficient cell lysis and neutralization each and every time.
Insufficient Mixing
Correct Mixing
Insufficient Mixing
Correct Mixing
Plasmid DNA Midi and Maxi Kits Specifications
Specifications
Cat. # PIF025 PIFE25 PI025 PIE25 PM010/25 PME10/25
Format
Anion-Exchange
Gravity Flow
Anion-
Exchange
Gravity Flow
Anion-Exchange
Gravity Flow
Anion-
Exchange
Gravity Flow
Anion-Exchange
Gravity Flow
Anion-
Exchange
Gravity Flow
Binding Capacity 500 µg 500 µg 800 µg 800 µg 1.5 mg 1.5 mg
Filter Column Yes Yes N/A N/A N/A N/A
Endotoxin Level <0.15 EU/µg <0.10 EU/µg <0.15 EU/µg <0.10 EU/µg <0.15 EU/µg <0.10 EU/µg
Culture Input 50-150 ml 50-150 ml 50-300 ml 50-300 ml 200-1600 ml 200-1600 ml
Plasmid Size 1-20 kb 1-20 kb 1-20 kb 1-20 kb 1-20 kb 1-20 kb
Typical Yield 200-500 µg 200-500 µg 200-800 µg 200-800 µg 1-1.5 mg 1-1.5 mg
Elution Volume 500 µl-2 ml 500 µl-2 ml 500 µl-2 ml 500 µl-2 ml 500 µl-2 ml 500 µl-2 ml
Operation Time <80 minutes <80 minutes <120 minutes <120 minutes <180 minutes <180 minutes
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