Sequencing Services From MWG Biotech

In our laboratory we require up to 30 sequencing reactions each month. This number is too low to justify the purchase of a sequencing machine. So, we made the decision not to sequence our clones ourselves but rather to look for a good, reliable partner who can do this expensive and time-consuming task for us. There were several requirements that the sequence facility had to meet, such as reliability, ability to produce long read-throughs, fair prices and high production speed.

We have sent clones to several companies offering sequencing services. The quality of the resulting sequences varied considerably. In the beginning of 2006, we started to send sequences to MWG Biotech AG in Germany. They have a somewhat elaborate ordering system which uses barcodes and on-line order forms. Via their website, you order barcode stickers; the stickers are free of charge and you can order hundreds at a time, ensuring that you never run out. You have to enter the number from the sticker in the on-line form together with the template name and type of DNA (plasmid or PCR product). Then place the sticker on the tube containing the template DNA. You need to use a single tube for each sequencing reaction. Finally, send the tube to the sequence facility in Germany. It takes some getting used to, but it works very well. You can choose between sequencing primers from MWG Biotech or supply one of your own. Rather than sending a tube with primer, it is also possible to supply a primer sequence which is produced at the sequence facility and used for sequencing. For plasmids, they recommend sending 1-2 µg of purified, air-dried DNA. Send 20 ng DNA/100 bases for PCR fragments (also purified and air-dried). Since purity can be a problem for PCR-products we always send plasmid DNA. Our routine protocol is to purify PCR products from an agarose gel, ligate the product into a pGEM®-T easy vector (Promega, cat. no. A1360) and transfer to XL2-Blue competent cells (Stratagene, cat no. 200150). The plasmids are purified using the Qiaprep Spin Miniprep Kit (Qiagen, cat. no. 27106). Finally, the samples are dried using a speedvac and shipped to MWG Biotech.

Results are usually returned to us in 4-5 days via e-mail. The email contains the sequence as text, but via a provided link, you can download the chromatogram data. At this point, we have sent more than 350 clones with greater than 95% generating successful sequencing reactions. Reactions are considered successful if they have a readable length of at least 900 bases, but read-lengths are usually more than 1000 bases. This is far better than the results we usually got from other sequencing facilities (up to 600-700 bases). For a single sequence, we pay 11.70 Euros. This is quite expensive but having to sequence clones more than once is usually more expensive. Since we only have a few clones to sequence each month, the cheaper 96-well ordering system is not an option. MWG Biotech also offers a prepaid sequencing system which reduces the cost approximately 2 Euros per sequence. In our experience, MWG Biotech offers the best sequencing services with the highest success rate.