SupremeRun Plate Service

Sample preparation & sumbission for the SupremeRun 96 service

• Please use fully skirted PCR plates  
• Template concentration must be normalised across the plate
• PCR product size should not vary by more than a factor of 3
• Samples should be sent liquid in a total volume of 15 µl
• Seal your plates using 8-cap strips to prevent material loss
• Stick the SupremeRun 96 plate barcode on the plate edge
• Label your enclosed primer with our free Primer Barcodes
• Ship samples at ambient temperature to our sequencing lab

Quantify the template concentration via agarose gel or a photometer to ensure accurate results.

Benefit from the greatest flexibility to handle your sequencing primers.

Thanks to our DNA synthesis lab, bioinformatic know how and internal IT solutions you can choose how to provide us with your sequencing primers:

• Select from standard primers on stock
  Select and manage your favourite standard primers from more than 80 different validated standard primers
• Order specific sequencing primers
  Specific primers can be ordered directly with your sequencing order. These primers are stored for 4 weeks and can be re-used during this time
• Enclose your own sequencing primers
  You can send us your sequencing primers in 1.5ml tubes along with your samples by using our Primer Barcodes

Please consider optimal primer conditions, concentration and volume as described below.

Optimum primer conditions:

• Primers must not contain phosphorylation or fluorescent dyes
• The optimum primer length is between 16-25 bases
• The primer melting temperature (Tm) should be 50 - 62°C
• The GC content of the primer should be 35-60%
• Ideally one G or C should be located at the 3' primer end
• The number of 3' Gs or Cs should not exceed 2 Gs or Cs
• If possible, avoid >3 identical bases in a row in the sequence

Primer concentration and volume:

• Exactly 10 pmol/µl primer concentration is required per sequencing reaction
• Each primer must have a total volume of 15 µl (double distilled water or 5mM Tris-HCl); 5 µl of primer volume is required for every additional sequencing reaction
• Concentration of primers with wobble bases must be calculated according to the following formula: nX x ConcPrimer

n = number of bases within a wobble according to IUPC code, X = number of wobbles within the primer sequence. [e.g. 1 V (AGC) = 3¹ x 10 pmol/µl; 2 V (AGC) (AGC) = 3² x 10 pmol/µl]

DNA and primer storage:

• Storage of template DNA for 4 weeks
• Storage of synthesised primers for 4 weeks free of charge
• Storage of synthesised primers for one year at additional cost
• Storage of enclosed primers for 10 days free of charge

Data storage:

• Secure online archive of all selected sequence data for 6 months

Definition of technical failure

A technical failure is defined as a chemical or technical (IT, machines) defect in our lab. In order to ensure that the sequencing quality is not affected by a technical failure, an internal quality is run with each plate (for that purpose well H12 should be kept empty, otherwise a quality control is not possible). In addition to that, a sophisticated software is analysing each plate considering parameters like quality values (QV) and reading lengths. If these parameters fall below a certain percentage across the plate an additional manual check is performed. If a technical failure is confirmed by this manual check, the plate is getting re-sequenced.