Frequently Asked Questions About Our Products And Services

All prepaid barcode labels, kits, and coupon codes remain valid for five years from the date of ordering, after which they expire.

You can opt to receive an email notification before any of your prepaid Sanger products expire by specifying it in your personal preferences.

Tube Labels are free barcode labels used for error free labelling and identification of your samples in tubes for all kind of Sequencing services. They allow fast processing your sequencing order. You can track the usage status, the usage date as well as the user of the lables under "May Sanger Kits & Labels".

Tube Labels can be ordered online and are free of charge.

If you send your own primers for sequencing, use the yellow primer barcodes to identify them.

An overview of which labels were used when and by whom can be found under "My Sanger Kits & Labels".

Primer barcodes can be ordered online for free.

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We will keep your data 6 months after finishing your reads in your personal account. During this time, you can download and save your data.

For all services, DNA samples are stored for 4 weeks.

Synthesised primers are stored for 4 weeks free of charge.

Storage of synthesised primers for one year is available at additional cost.

Enclosed primers are stored for 10 days free of charge.

Storage of enclosed primers for 6 months is available at additional cost.

During this time it is possible to use the DNA and primer for further reactions.

Premixed samples (DNA mixed with the primer) and pre-cycled samples for the Ready2Load service will be discarded after the order is finished and the data submitted.

Yes, we accept templates with more difficult sequence structures. 

Please contact our Customer Support to get the best recommendation for your request.

Overview of delivery times for the majority of our Sanger services:

 
Please note:

Specified delivery times are mainly depending on sample arrival time in the lab.

For many European cities we are providing coordinated pick-up and transport services using Eurofins DropBoxes ensuring on time delivery in the lab.

However, these logistics services may also be subject to force majeure or unpredictable changes caused by e.g. strikes, accidents, storms, floods. In such events, Eurofins Genomics cannot guarantee the stated data delivery time and is not responsible for delays caused by logistics services.

Additionally, our general terms and conditions (GTCs) apply.

Please use our online shop for every Sanger Sequencing product and service. In the order menu you find all links to the respective sequencing order pages. 

You will find all necessary information regarding sample preparation for each sequencing service under in the sample submission tab of the respective service.

Eurofins Genomics has already installed hundreds of DropBoxes in Europe for free sample pick-up. Your nearest DropBox location with pick-up time is displayed in your account when you are logged in.

In case there is no DropBox near you, you can of course send your samples by post or arrange for them to be delivered by another delivery company. Please send Sanger sequencing samples to the addresss which is shown to you during checkout.

Yes, indepenent on the sample type, more reactions per sample can be specified. Just follow the instruction on the order page. 

Up to 15 plates can be uploaded / specified at once and saved in you cart. There is no limitation on the number of plates per order you can checkout.

You can order new sequencing reactions from stored samples within 4 weeks no matter if you have sent the samples in tubes or plates. 

If you have sent the samples in tube format, just use the same order page and type in the initials of the respective sample. The system prepopulates all stored samples with these initials.

If you want to re-order reactions from a few samples originally sent in plates, you find the option "Re-Order from stored plates". Select the stored plate and specify the new reactions for the respective samples.

Under "My samples & primers" you can check the expiration date of your samples and primers.

With prepaid barcodes, you can order additional services such as primer synthesis DNA mini preparations. You will be billed separately for this.

You will find all information about sample format, template and primer concentration as well as shipping information for each of our sequencing services in the respective "Sample Submission" tab.

Please measure the OD with a spectrophotometer at 260 nm and 280 nm. The OD 260 values should be in the range of 0.05 to 0.8 to give reproducible and reliable results. The OD 260/280 ratio should be about 1.8. Values lower than 1.6 and higher than 2.0 indicate contaminants in the sample that interfere with the determination of the concentration and might inhibit the sequencing reaction.

If possible, we recommend measuring the ODs at 230 nm and 320 nm, too. A high value at OD 320 indicates a contaminant. This should be ideally 0.0. The OD 230/260 ratio should be lower than 0.6.

We also recommend ensuring the quality of your DNA by running your sample on an agarose gel.

For plasmids, we recommend the use of commercially available plasmid mini scale preparation kits employing spin columns. From our experience, repeating the washing step will improve the quality of the DNA and therefore improve the sequencing results. Please do not send DNA prepared with alkaline lysis methods or with the boiling method (Holmes and Quigley, 1981) without column purification. For PCR products please use commercially available PCR purification spin column kits.

No, please don't.
EDTA binds bivalent cations such as Mg²⁺ that are essential for the Taq polymerase. Your DNA is stable in double distilled water or Tris-HCl at room temperature for several days or even longer if it is dried down.

This is not necessary; DNA is stable in water at room temperature for several days. Just send your samples either in Tris-HCl, distilled water or air dried. Please, do not use EDTA as this will inhibit the Taq polymerase.

Yes, for all sequencing services we offer template preparation and PCR product purification as an additional service.

Successful isolation of plasmid DNA is possible from most E.coli strains, though the strain which is used can have a significant influence on the quality of the purified DNA. Host strains such as DH10b, DH5 alpha, XL10 Gold and TOP10 normally result in high quality DNA in combination with many commercially available plasmid DNA isolation kits and are therefore ideal for the propagation of plasmids to be sequenced. Lower quality DNA is derived from strains producing large amounts of carbohydrates, which are released during lysis and inhibit enzyme activity. Examples are HB101 and its derivatives such as TG1 and the JM100 series. Also strains with medium or high levels of endonuclease activity like HB101, JM101 or BL21 generate DNA of lower quality, hence a proteinase K treatment should be considered.

Yes, primers and dNTPs must be removed from the product. The primers would produce a mixed sequence (5% contamination with a primer results in unreadable mixed sequences!) and the dNTPs would interfere with the dNTP/ddNTP concentration in the sequencing reaction. We recommend purification by using commercially available PCR spin column kits. Alternatively we can perform the PCR product purification for you as additional service.

If you want to employ one of the PCR primers in the sequencing reaction you must elute the product of interest from an agarose gel. Otherwise, a mixed sequence will be produced as the primer is most likely to bind both the specific and unspecific products. If you employ a specific nested primer in the sequencing reaction, it might work. However, producing only one product in the PCR reaction is the best guarantee for a good sequence. In almost all cases, improving the PCR conditions (specific primer design, higher annealing temperature and/or lower primer concentration) helps to avoid unspecific products.

80 different standard primers are available for your DNA sequencing order. for free Under "My samples & primers" you see the list of standard primers.

Yes of course. Just specify your enclosed primers in your sequencing order with the primer name. The primers should be sent in a solution and concentration specified in the respective "Sample Submission" pages. Please use a primer barcode to identify your primer.

Yes, we can synthesise your sequencing primer in our oligo production. Just specify your primer for synthesis in your sequencing order with the respective name and sequence. 

Please find information about primer format, primer concentration and primer shipment information for each of our sequencing services in the respective "Sample Submission" tabs.

The primer synthesis along with Eurofins sequencing services can be payed with prepaid barcodes.

DNA base call identification is based on the nomenclature system issued by the International Union of Pure and Applied Chemistry (IUPAC). At positions of ambiguity, the following IUPAC codes are used:

Eurofins Genomics is using a quality base calling software which examines the peaks around each base call to assign a quality score (Q) to each base call. Quality scores (Q) range from 4 to about 60, with higher values corresponding to higher quality. The quality scores are logarithmically linked to error probabilities, as shown in the following table:

For all our Sanger Sequencing Services we are using the cycle sequencing technology (dideoxy chain termination / cycle sequencing) on ABI 3730XL sequencing machines.

Cycle sequencing is a modification of the traditional Sanger sequencing method. The components are DNA, primer, heat resistant DNA polymerase, 4 dNTPs, 4 ddNTPs (dideoxy terminator nucleotides) fluorescently labelled with four different dyes and buffer containing Mg⁺⁺ and K⁺. The single primer binds to the complementary DNA strand and is extended in a linear mode.
This extension continues until by chance and depending on the complementary base a particular ddNTP is incorporated. Because of the latter's dideoxy-configuration the polymerase cannot add any other base to this fragment and the extension is terminated. Thus at the end of the selected number of cycles, numerous fragments with different lengths and one labelled nucleotide at the end are generated.

Stoichiometric manipulation of the reaction components ensure that the fragments of every possible length starting from n+1 say 2000 bases are generated with n being the number of bases in the primer. The key difference between the traditional Sanger method and cycle sequencing is the employment of a thermo stable DNA polymerase. The advantage of using such a polymerase, is that the sequencing reaction can be repeated over and over again in the same tube by heating the mixture to denature the DNA and then allowing it to cool down to anneal the primers and polymerise new strands. Therefore less template DNA is needed than for conventional sequencing reactions.

After a post sequencing reaction cleanup, the samples are electro kinetically injected into the array of 96-capillary sequencers. The negatively charged fragments migrate toward the anode by size, the smallest ones move fastest. Their tagged ddNTP terminators can be read as the fragment's base sequence. A laser beam excites these dye molecules as the fragments reach a detection window, producing fluorescent signals that are collected from all 96-capillaries at once, spectrally separated and focused onto a CCD (charge coupled device) camera. Sophisticated optical and electronic devices produce a colour readout that is translated with the help of sequence analysis software into a sequence.

The quality report is the trace file in *.pdf format in which the quality value of each single base is shown in colour code below each single peak. Different colours represent the four specified quality ranges.

Under the conditions listed below, only simplified test reports may be provided.

Subject: Simplified test reports according to DIN EN ISO 170252018, (Section 7.8)

Testing Laboratory (Accreditation): Eurofins Genomics Europe Applied Genomics GmbH (D-PL-13372, ) Eurofins Genomics Europe Sequencing GmbH (D-PL-17038)

For accredited examinations at these two companies, test results may only be reported in simplified form, i. Requirements of the standard DIN EN ISO 17025: 2018 (section 7.8) are not fully applicable to this type of transfer of results.

These results can therefore also be reported as short test reports, electronically generated result reports, Excel spreadsheets or as result files.

 

If you have any questions, please contact the Customer Care Team.

There are many softwares on the market which you can use for testing or buying.

For your convenience you can download and install our GATCViewer software for free.

In order to ensure that the sequencing quality is not affected by a technical failure, an internal quality control is performed with each sequencing run. In addition to that, a sophisticated software is analysing each plate considering parameters like quality values (QV) and reading lengths.

If you see "DONE_QV_NOT_MET" as a result of your sequencing reaction, it means that the internal quality control has passed, but some sample quality values have not been met. (e.g. sample or primer concentration or volume, sequence design or structure etc.).

There are pros and cons of every sequencing method. Sanger sequencing and Oxford Nanopore sequencing (ONT) are both methods for determining the sequence of nucleotides in a piece of DNA. Typically Sanger is consider the most accurate method for short-read sequencing and NGS is better for long-read sequencing.

Overall, the choice between Sanger sequencing and ONT will depend on the specific needs of the application. Both methods have their strengths and limitations, and the appropriate method will depend on factors such as the length and quality of the DNA sample, the desired level of accuracy, and the cost and availability of the necessary equipment.

It really depends on the sample quality. We cannot guarantee the level of coverage at this time. A good sample submitted properly will typically yields hundreds or even thousands of sequencing reads. Consensus coverage depends on how many reads are full-length plasmids and how many, if any, degraded.

A coverage exceeding approximately 20x indicates a highly accurate consensus sequence.

A coverage below 20x can include inaccurate assembled plasmid sequences. The most common reasons for low quality coverage assemblies are listed in ‘Troubleshooting guide for failed samples’.

Any number of samples is welcome. There are no minimums or limitations.

You can send it and we can sequence it, but we cannot predict or promise the analysis outcome. The customer would take on the risk of these orders.

Great news! Yes we do. All information about our prepaid solution can be found here >>

Unfortunately not. Currently, we can only accept plasmids. However, we do offer a special service for amplicons and large DNA inserts, which is available for your use. Learn more >>

Unfortunately not. This barcode is required for our internal lab processes and is essential in ensuring that we can process your samples as efficiently as possible. Thank you for your cooperation!

Yes of course.

If you don't know if there is a dropbox close by, use our dropbox location page to find out.Please click here >>

Yes, FASTQ files are delivered for every project.

There are several reasons why your sample might have failed:

1. The starting material did not consist of circular DNA.
   In our library preparation protocol, circular DNA is required as the starting material. If linear DNA is provided, it will result in sample fragmentation and subsequently lead to poor or no results.
   
   
2. Insufficient DNA concentration in prepared samples.
   The primary reason for this issue is the use of a Nanodrop device to measure DNA concentration. We strongly recommend employing a Qubit or a similar method instead.
   
   
3. Presence of mixed plasmid species and/or fragmented genomic DNA or fragmented plasmids within the samples.
   You may detect signs of this failure mode through a wide range of read lengths reported in the raw read length histogram.
4. Contaminants in your samples like Ethanol. For more info please click here: https://community.nanoporetech.com/contaminants

To interpret your results you can also have a look at our troubleshooting guide. Click here >>

Please send your samples in nuclease-free water or elution buffer.

Please do not use a buffer containing EDTA.

For each run, we conduct multiple quality control (QC) measurements to assess the success of the library preparation and sequencing. If all the measurements indicate satisfactory results, we will not reprocess samples that have failed.

We will sequence your plasmids typically within 1 business day.

For plasmids > 25 kb please except a turnaround time of up to 5 business days.

The prevalent error modes in Oxford Nanopore sequencing include deletions within homopolymer stretches, errors occurring at the central position of the Dcm methylation site CCTGG or CCAGG, and errors at the Dam methylation site GATC.

So please handle these regions with special care.

Unfortunately not. The protocol used for the high-throughput Whole Plasmid Sequencing service does not allow single-stranded DNA.

But of course we can sequence your ssDNA molecules with our custom workflow. More information can be found here >>

• In order to offer our competitively priced product, we pool different samples on flow cell runs and re-use flow cells by washing them between runs. Both procedures are industry standard and performed according to the manufacturer’s specifications.  
• Barcoding and pooling samples on a single flow cell reduces cost and maximizes usage efficiency of the pores. However, nanopore sequencing has a higher error rate compared to “traditional” short-read next-generation sequencing, which can lead to misreadings in the barcode sequence and incorrect sample assignment/de-multiplexing at a rate of up to 0.05%. This issue is known as index hopping. 
• The flow cell washing procedure is very effective with only up to 0.1% of any previously loaded sample contaminating a subsequent run (known as carry-over contamination), nevertheless you might experience individual contaminating reads within your raw data.
• However, for both index hopping and carry-over it is important to note that a contamination of this level is unlikely to impact the final sequence assembly as several fine-tuned sequence cleaning and filtering steps are applied, provided your sample meets our quality and quantity standards and therefore yields sufficient numbers of high-quality plasmid reads. It is important to note, that the issue is most prominent in low coverage assemblies (see ‘What is the coverage?’).
• In the case that customer samples do not adhere to our sample specifications, the samples are sequenced on the customer’s own risk, and we cannot assume any liability for potential damages caused.
• If you’re concerned about contaminations and most importantly about reads from your samples appearing in other customer’s raw data, we offer the possibility to use our premium ONT services which include usage of a dedicated full flow cell for individual customers (learn more >>).

Here are a few kits:

VWR Plasmid Miniprep Kit II

QIAGEN Plasmid Plus Midi Kit 

peqGOLD Plasmid Miniprep Kit II

ZymoPURE II Pure Midiprep Kit 

For really large plasmids different kits are required. For example ZymoPUREII or NucleoBond.