How tos and FAQs

General rules of sample preparation

If you order sequencing or fragment analysis in tubes, please use preferably 0.2 ml strips for your samples. You can use other tube types or 96-well PCR plates too.
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StandardSeq – Preparation of samples

Sample is a template mixed together with sequencing primer in a single test tube / well or template only if you wish to use some of our universal primers. Template is a PCR product or plasmid.
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CleanSeq – Preparation of samples

Sample is an unpurified PCR product (aliquot of the PCR reaction) or other suitable template, amenable to enzymatic cleanup. You must also supply sequencing primer(s) or select some of our universal primers.
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HairpinSeq – Preparation of samples

Sample is typically a plasmid with a cloned hairpin or GC-rich stretch in the region of interest or PCR product. You must also supply sequencing primers (forward and reverse) or select some of our universal primers.
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ReadySeq – Preparation of samples

Sample is a sequencing reaction prepared by using the BigDye Terminator v3.1 Cycle Sequencing kit (Applied Biosystems), cleaned and finally dissolved in at least 15 μl of Hi-Di formamide (Applied Biosystems).
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What is the difference between sequencing protocols you offer?

StandardSeq is the basic protocol, we sequence your template. CleanSeq is in fact StandardSeq with template cleanup performed beforehand. HairpinSeq is a special protocol ...
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Fragment analysis – Preparation of samples

The sample is typically a PCR product or a mixture of PCR products which are fluorescently labelled. The PCR products do not have to be purified, you only need to send the aliquot of the PCR reaction.
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Can I copy-paste into the order form?

Definitely yes. Create (in Excel, for example) a table with three columns - Sample Name, Primer Sequence and Template Size (applies to StandardSeq, CleanSeq, ReadySeq). For HairpinSeq the table must have four columns - Sample Name, Primer1 Sequence, Primer2 Sequence and Template Size.
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Universal primers

The following universal primers are supplied by us free of charge.
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My own primers

When designing custom primers, please remember that properly designed primer is crucial for sequencing...
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How should I specify primer sequences?

Please use ACGTN symbols for the primer sequences, IUPAC codes (BDHRYKMSWV) for degenerate bases and “I” for Inosine. If there are...
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I want to use my own primer which you use as a universal primer, how can I do this?

If you want to use for example your own M13F primer, you have two choices. Either you do not enter primer information in the order form and only add it to your template (StandardSeq protocol), or you specify this primer in My Primers...
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How should I specify the template size?

If the template is a PCR product, enter its size. If the template is plasmid with a cloned insert, enter the total size of the template (plasmid plus insert). This field is optional for ReadySeq.
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There is a limit on template length in the order form for Sanger sequencing. Why?

This is correct, the template size must be between 100 bp and 20.000 bp. It is quite unusual to sequence templates shorter than 100 bp. Assuming your primer has 20 bases ...
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How do I ship samples?

We recommend you insert samples into a quick-closing plastic bag. Fragment analysis samples can be (not necessarily) wrapped up in aluminium foil to prevent light access.
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Subscribing sequencing services

Click on link New Prepayment in your account and complete the form to subscribe to our services. The amount to be prepaid is given without VAT, the system will add VAT automatically according to applicable regulations. Once you have confirmed your order, an advance payment request will be issued automatically which you can use to ...
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Other questions?

Do not hesitate to contact us
We will be glad to help you.

Richard Nádvorník
+420 602 102 790
info@seqme.eu

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