w6.0.20 | c6.0.57.18
PROD | u6.1.6
Header Image service-corner
 

 

print content
increase font
decrease font

Frequently Asked Questions About Our Products And Services

Custom DNA Sequencing

Products & Services

What is the Tube Label - when / why do I need it?

Tube Labels are used for error free labelling of your sample tubes and enable less paperwork.

They allow fast and convenient processing of your samples for all kind of Custom Sequencing Services in tube format, available on our Ecom system.
They are also used to recognise each customer identity.
You can order these Tube Labels free of charge in your online account.

What are Prepaid Barcode Labels?

Prepaid Barcode Labels can be ordered in sets of 50 for our Value Read sequencing service in tube format.

Prepaid Barcode Labels are available in batches of 50 and offer the following features:

  • Easy ordering and up-to-date tracking of labels online
  • Flexible usage to facilitate sharing among colleagues
  • Simple labelling your Value Read sample tubes
  • Convenient payment of your primer synthesis

What are PlateSeq Kits resp. PlateSeq Coupons?

PlateSeq Kits and PlateSeq Coupons are the prepaid solution for our PlateSeq service.

Each PlateSeq Kit comes with a 96well plate and the appropriate seals in a metallic sample box. For each template type we offer the perfect PlateSeq Kit.

PlateSeq Coupons are code numbers provided electronically in your online account. You can use them:

  • For a second prepaid sequencing run in addition to one of our PlateSeq Kits (e.g. forward & reverse sequencing reads).
  • As alternative to the PlateSeq Kits for plasmid DNA, purified PCR products or premixed samples for immediate use.

Prepaid solutions reduce your administrative efforts and additionally help you to save your budget as we offer a lower price compared to the non-prepaid PlateSeq service.

How long will my sequencing data be stored?

We will keep your data 100 days after finishing your reads in your personal account. During this time, you can download and save your data.

How long do you store samples and primers?

For the Value Read service in tubes we store your custom synthesised primers for 6 weeks. Your DNA samples and enclosed primers will be discarded three days after the order is closed.
For the Sequencing Service à la Carte and for the PlateSeq Service we store your provided samples and primers for 6 weeks after arrival.
For the Walking Services we store your samples and the primers for 3 months. For our GLP - Compliant Sequencing service we keep your samples and primers for minimum one year.

Can I send shRNA constructs or other DNA templates with probable secondary structures?

Yes, please choose our Sequencing Service à la Carte to place your order.

Can I send Phage DNA?

Yes, please choose our Sequencing Service à la Carte to place your order.

Can I send genomic DNA?

Yes, please choose our Sequencing Service à la Carte in combination with our PCR amplification service to place your order.

What is the difference between the primer walking service and the GLP-compliant sequencing service?

The sequencing process itself for GLP-compliant sequencing by primer walking (sequence analysis compliant with Good Laboratory Practice) does not differ from the sequencing process for double strand sequencing by primer walking.

The difference makes the comprehensive GLP-compliant project documentation according to FDA guidelines.

How to order

How can I order your DNA sequencing service?

For your convenience, please use our Ecommerce (Ecom) system to place your order. 
If you cannot order online, please download one of our order forms.
Please be aware that the Value Read Tube and the Ready2Load Sequencing services can only be ordered online.
You will find all necessary information regarding sample preparation for each sequencing service under "Sample Submission".

Where should I send my sequencing samples?

For Europe please send to:
Germany
Eurofins Genomics  
Sequencing Department
Anzinger Str. 7a
85560 Ebersberg

For customers in the following countries, please send your samples to:
United Kingdom
Eurofins Genomics
i54 Business Park
Valiant Way
Wolverhampton
WV9 5GB

France
Eurofins Genomics 
ZA de Courtaboeuf
9 avenue de la Laponie
91978 Les Ulis

Italy
Eurofins Genomics
c/o Regusa
Via Senigallia 18/2
Torre A
20161 Milano

How do I provide a reference sequence?

Please paste your reference sequence into the provided field in the respective order form on our Ecom system. Alternatively you can send your sequence information together with your filled order form (downloadable from our website) by e-mail to: sequencing-eu@eurofins.com.

Can I order more than one read per sample?

Yes, of course.
With our Single or Double Strand Walking services we cover the complete sequence of your plasmid insert or PCR product and we manage the design for the walking primers too.

For the Sequencing Service à la Carte you just need to send us the required amount of DNA or the clone in one tube, and specify the number of reactions in your order form.

If you want to send us a clone or an unpurified PCR product for more than one reaction using our Value Read Tube service, just place a sample name into the order form and mark this tube individually. In addition please include the corresponding amount of Barcode Labels - one for each sequencing reaction - in your sample package. Transfer the final 3 letters of each of your barcodes into the designated fields within the order template and send your sample tube together with the Barcode Labels to us.
If you send us plasmid DNA or purified PCR products for our Value Read Tube service we need one tube per reaction as this service is highly automated.

How many plates per order can I send?

A total of 8 plates per order can be sent for sequencing.

Can I re-order sequencing runs?

You can order new sequence runs without sending new sample material by taking advantage of our free template & primer storage service for

  • PlateSeq Service
  • Sequencing Service à la Carte
  • Primer Walking services 

You can order either a new sequence read on a stored DNA template with a new primer or on a new DNA template with a stored primer.

For our PlateSeq service you have not only the possibility to order the re-sequencing of complete plates but to re-order new sequence reads of single wells with different primers.

Sample Preparation

How should I send my sequencing samples?

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.

How should I determine the concentration of my samples?

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.

Which purification method should I use?

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.

Can I send my DNA samples in Tris-EDTA (TE)?

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.

Should I submit my DNA samples cooled?

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.

Can I send unpurified PCR products or clones?

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

Simply send us your un-purified PCR product at room temperature.

Which E.coli strain should I use?

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.

Do I need to clean up my PCR product if there is only one band visible?

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.

I see unspecific bands in my PCR reaction. Do I need to gel purify my product?

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.

Primers & More

Which standard primers do you offer?

More than 70 different standard vecotor primers are available for your DNA sequencing order. Please click here to find the complete list of our standard primers which are available free of charge.

Can I send my own primers for sequencing ?

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

Can you synthesise a primer for me? How do I order the synthesis?

Yes, we can synthesise your sequencing primer in our proprietary DNA synthesis facility. Just specify your custom synthesised primer in your sequencing order with the respective name and sequence. Please note, that we keep these primers for 6 weeks for re-order possibilities and discard them afterwards.

How should I send my primers and how much primer do you need?

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

Technical Questions

What is a BLASTx analysis? What results will I receive?

BLAST (Basic Local Alignment Search Tool) is the umbrella term for a vast range of world wide used programs to analyse biological sequence data.
BLASTx is used to compare a newly determined DNA sequence against already existing sequences in the NCBI non redundant protein database. This service is used to find a potential translation of an unknown nucleotide sequence.
In your account you will get a text file with the original BLASTx output as well as a report as HTML with the best BLASTx hits. Both files are stored in your account for 100 days and can be downloaded during this time.

What are IUPAC bases? Which results will I receive?

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:

G/T = K C/T = Y A/C/G = V
A/G = R A/C = M C/T/G/ = B
G/C = S A/G/T = D A/C/G/T = N
A/T = W A/C/T = H

What is the plate result summary?

The plate result summary is the result overview of your sequenced samples per plate.
It contains the names of each sample, the read lengths of each sample, the quality clipping positions and the overall quality value of the sequence.

What are Q20 (Q30 / Q40) bases?

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:

Quality ScoreProbability of a wrong base callAccuracy of a base call
Q 10 1 in 10 90 %
Q 20 1 in 100 99 %
Q 30 1 in 1.000 99.9 %
Q 40 1 in 10.000 99.99 %
Q 50 1 in 100.000 99.999 %

Which sequencing method do you use?

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

Could you explain the cycle sequencing technique?

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.

What is the quality report?

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.

Scroll to top ^^