Services | Midipreps (+DNA Sequencing)

Safety

• As with all our services where we are accepting live bacterial material, please note that we can only accept samples that have been classified as biohazard containment level 1 according to the UK Genetically Modified Organisms (Contained Use) Regulations and ammendments.
• For more information on these regulations, please see the HSE website.
• We may require proof of containment level to be supplied.

Having Your Midiprep Sequenced

If you would like us to sequence your completed midiprep, simply click "Midiprep + DNA Sequencing" above.

During the submission process, we will request additional information that will allow our facility to carry out this sequencing service for you including:

• Information regarding your primer
  
  • We have a number of Standard Primers available
  • Or you can provide your own at the time that you submit your miniprep sample
• Whether your template is known to be GC-Rich

Please contact us if anything is unclear!

Potential Yield of Plasmid DNA

• Please note that the quantity of plasmid recovered from a midi-prep is dependent on many factors and not just the size of the pellet or how cloudy the culture looks
• Plasmid Copy Number Considerations
  
  • High Copy Plasmids
    
    • We have found that the yield for a high copy number plasmid can be around 0.25mg total DNA.
    •  In our experience, we have also found that isolation of DNA from midi-preps of high copy plasmids grown in a volume of 200ml routinely overload the columns we use and can result in a low purity product thus we discourage customers from proceeding with their midi-prep in this manner. 
  • Low Copy Plasmids
    
    • When using low copy plasmids, please ensure that you have identified the optimal strain to use for plasmid growth prior to submitting your cultures.   

• ^Augmented Protocol: If your template has known secondary structure or a high GC content, you can select our augmented protocol - £0.50 will be applied per reaction to facilitate this.

Do you routinely need a large number of midipreps prepared and sequenced?

Why not contact our staff to learn about savings options?

Our pre-payment option will help you save more per reaction!

Primer

• Standard primers offered by MRC PPU DNA Sequencing & Services are free!
• Custom primer synthesis will reflect the size of the primer – the cost to the customer is £0.50 per base

Learn more about all our pricing and turnaround.

Please note that prices are exclusive of VAT, which customers outside The University of Dundee will be charged at the appropriate rate (currently 20%) on all products and that all sales are subject to MRC PPU DNA Sequencing and Services' Terms and Conditions.

What customers can provide:

We are happy to be flexible regarding receipt of materials to suit your needs!

• Plasmid DNA - Please review our Complete Midi-Prep Service below
• Bacterial colonies on a plate - Please review the information provided below
• Inoculated cultures to be grown - Please review the information provided below
• Fully-grown Culture/Spun-down pellets - Please review the information provided below

Labelling Samples

• When sending multiple midi-preps, please provide your samples in tubes labeled sequentially from "1" onwards
• As customers’ samples will be provided back in much smaller tubes than the culture tubes, excess text will hamper accurate transfer of sample name from one tube to another

E. coli Strains

• We have found that DNA isolated from those E. coli strains in which the Endonuclease A has been mutated, results in purification of DNA that is more fully intact (non-degraded)
  
  • Please consider this should you choose to send us starter cultures/spun down bacterial pellets
• Please note we only use the DH5alpha E. coli strain in-house

Media & Growth Considerations

• When growing your cultures, please use only LB broth
• Using too large a volume, an inappropriate growth media (ie. TB broth) or growing for too long results in a lot of bacteria but not much plasmid
  
  • Ultimately, excess bacterial cell material can clog up the clearing filter and leads to loss of your plasmid material
• Media Volumes:
  
  • 100ml recommended
• Time Considerations: We have found that conditions that favour plasmid isolation include those where growth of the culture does not exceed 16 hours

Bacterial Colonies on a Plate

• After you have transformed your plasmid(s) of interest, you can send us your plate(s) and our facility will:
  
  1. Select 1 colony per plasmid 
  2. Grow midi-prep cultures
     
     • Please inform staff about antibiotic resistance for each plasmid
  3. Isolate DNA
  4. Quantitate DNA concentration

Inoculated Culture To Be Grown

• Starter Culture: Please send your starter culture (5ml) together with details of the volume of culture and antibiotic to use
• Antibiotic: As standard we have -
  
  • LB + Ampicillin
  • LB + Kanamycin
  • If you require a different antibiotic, we ask you to please grow your own culture and provide these to the DNA Sequencing and Services facility
• Our facility can also isolate DNA directly from a culture you have begun growing

Fully Grown Culture/Spun-Down Pellets (Centrifuged Samples)

• Our facility can also isolate DNA directly from a culture you have already grown from:
  
  • A fully grown culture
  • Spun-Down Pellet
• To get the best results, ensure that you drain off all the culture medium, since not doing so reduces the quality of the plasmid DNA produced.

DNA Isolation

• The preps we perform are Invitrogen Pure-Link high speed midi-preps.
• After prepping, the DNA is checked for quantity and quality on our nanodrop spectrophotometer and the concentration will be provided with the sample.

Complete Midi-Prep Service

Should you wish for MRC PPU DNA Sequencing and services to take care of your midi-prep from start to finish, our facility can easily do so. Please provide samples of DNA and we can take it from there! We don't stipulate an exact amount of DNA, but if you send us 20ul of your plasmid at 10ng/ul, that will be enough. We will:

1. Transform your DNA
2. Pick a single independent colony to grow a culture per plasmid
3. Isolate the DNA for you
4. Quantitate DNA concentration

Starting from an already grown culture, our facility can routinely turn samples around within one day of receipt. Additional time will be needed in the following cases:

• Submission of Starter Culture - Routinely 2 working day turnaround time due to additional required steps (amplification of bacteria/plasmid)
• Submission of Plasmid DNA - At least 3 working day turnaround time due to additional required steps (transformation, culture growth).

Shipping Details

• Labels
  
  • Please ensure that samples are clearly labelled so that we can identify them.
• Packing Materials
  
  • Eppendorf Tubes (if sending plasmid DNA to be transformed)
    
    • Please also ensure that your samples are well protected and packaged to meet Post-Office or courier regulations
      
      • A "Jiffy" style padded envelope for DNA samples in plastic tubes works well
      • We discourage use of regular envelopes as in the past, we have received envelopes with tears and/or crushed tubes in them
  • Agar Plates (if sending colonies to selected for growth)
    
    • Please also ensure that your samples are well protected and packaged to meet Post-Office or courier regulations
  • E. coli Pellets
    
    • None needed if hand delivering (internal customers)
    • Packing in dry ice within a polystyrene insulated container is the only packaging method we can accept for items sent by courier

Postage Costs

• Customers are responsible for paying all postage / carriage charges
  
  • For DNA samples or agar plates, normal first class post should be sufficient
    
    • As this does not always arrive as "next day post", we recommend that customers who have deadlines to meet send their samples by a guaranteed next day delivery service
• Insufficient postage provided on a package will cause a delay in use receiving the package
  
  • If MRC PPU DNA Sequencing and Services is then charged upon receipt, please note that we will need to pass this cost onto the customer