• The Value Read process is optimised for stab or glycerol cultures with the selected E.coli clones, purified plasmid DNA and unpurified or purified PCR products
• Use 1.5 ml tubes or 96well microtiter plates for your samples and primers. Please note that we require one tube per sequencing reaction
• For samples in tubes we ask to dissolve the DNA either in doubled distilled water or in 5 mM Tris-HCl (pH 8.0 – 9.0) in a total volume of 15 µl without using parafilm
• Plates may contain a mixture of plasmids and PCR products as long as no purification is needed
• For optimal results the concentration/amount should be normalised across the plate
• Primers may be arrayed in a separate primer plate according to the sample layout
• Plate position H12 should be kept free for internal quality control (optional)

Concentrations:

• Purified plasmid DNA:
  50-100 ng/µl in a total volume of 15 µl
• Purified PCR products:
  < 300 bp:            2 ng/µl in a total volume of 15 µl
  300-1000bp:      5 ng/µl in a total volume of 15 µl
  > 1000 bp:        10 ng/µl in a total volume of 15 µl
  The amount has to be doubled for unpurified products
  
• Primer Concentration
  2 pmol/µl, minimum volume: 15 µl (corresponds to an amount of 30 pmol)

Premixed Samples in Tubes or Plates:

• Purified plasmid DNA:
  50-100 ng/µl in a minimum volume of 15 µl incl. 15pmol primer
• Purified PCR products:
  < 300 bp:            2 ng/µl in a total of 15 µl incl. 15pmol primer
  300-1000bp:      5 ng/µl in a total volume of 15 µl incl. 15pmol primer
  > 1000 bp:        10 ng/µl in a total volume of 15 µl incl. 15pmol primer

• Use 1.5 ml tubes or 96well microtiter plates
• Use ABI Big Dye Terminator Kit (v3.1)
• Big Dye volume per reaction should not exceed 1 µl
• Total reaction volume of 10 to 20 µl
• Shipment of samples on dry ice is strongly recommended.

• It is possible to send either a stab or glycerol culture with the selected E. coli clone, the purified DNA, your unpurified or purified PCR product or genomic DNA
• Please use 1,5 ml tubes for your samples and primers
• We recommend to dissolve the DNA either in double distilled water or in 5 mM Tris-HCl (pH 8.0 – 9.0) without using Parafilm

Concentrations:

• Purified plasmid DNA:
  50-100 ng/µl, minimum 1 µg per reaction;
  Please send double amount for samples with hairpin- or secondary structures, high GC content or other difficult regions
• BAC-, PAC-, Cosmid-, Fosmid-DNA:
  200-1000 ng/µl, minimum 4-6 µg per reaction
• Purified PCR products:
  < 300 bp:            2 ng/µl in a minimum volume of 15 µl
  300-1000bp:      5 ng/µl in a minimum volume of 15 µl
  > 1000 bp:        10 ng/µl in a minimum volume of 15 µl
  The amount should be doubled for unpurified products
• Genomic DNA
  5-50 ng/µl, minimum amount is 200 ng per reaction
• Primer concentration
  2 pmol/µl, minimum volume per reaction is 10 µl (corresponds to an amount of 30 pmol)

• It is possible to send either a stab or glycerol culture with the selected E.coli clone, the purified plasmid DNA or your unpurified or purified PCR product
• Please use 1,5 ml tubes for your samples
• If available, please send a reference sequence in electronic form

Minimum concentration:

Plasmid DNA: 100 ng/µl
PCR products:  10 ng/µl


Minimum total amount:

• Single Strand Sequencing:
  Plasmid DNA: 1 µg/1 kb insert
  PCR products:  100 ng/1 kb PCR product

• Double Strand Sequencing:
  Plasmid DNA: 2 µg/1 kb insert
  PCR products:  200 ng/1 kb PCR product

• 4fold Coverage Sequencing:
  Plasmid DNA: 4 µg/1 kb insert
  PCR products:  600 ng/1 kb PCR product