Thanks to all of you who answered my question about cheap ways to score microsatellites, looks like they still have to invent a really cheap and precise device, anyway here they are: Hi Andrea, The spreadex system from Vhbio resolves to 2bp and scores 96 samples for c.£20. Unfortunately you'll only be able to score one loci at a time but it provides a relatively cheap solution with a small set up cost. You can stain the gels with Syber gold which is less time consuming than silver staining. The gels are fairly robust but you need a special rig and power pack to run them. The obvious alternative is a sequencer if you run your PCR products in a multiplex PCR etc. Our company charges £2.00 per run. For instance we use an eight primer multiplex PCR to genotype cherries which makes it a more cost effective solution. Best of luck. Jake Hi We use a Licor sequencer and we are very happy because the reagents and glass for gels are very cheap. We only have one banding colour but it works quit well and you can do a lot of PCR's on multipelx depending on mSats number of bp. Good luck Best regards Hello Dr. Verardi, I am responding to the email you sent to the EvolDir concerning scoring of microsatellite data. Our not-for-profit facility (in Canada) does this for academic labs at a very reasonable cost (on our ABI 3730 instruments)- what is your sample size and number of loci (are they multiplexed or pooled). I would happy to give you an estimate for this, if you decide not to purchase an instrument for your facility. However, I would recommend the ABI 3100 instrument as a reasonably-priced instrument to score fluorescently-labelled microsatellite fragments - it can run approx. 3 plates of 96 samples per day. Best Regards, Tara Paton Hi Andrea, One alternative would be to look for a used sequencing machine. It's likely that this may actually be just as, or more, expensive as sending samples out but you never know. Maybe something on e-bay or other used lab equipment sites. The only caution there is whether you can still obtain the necessary supplies to continue running the machine. Actually, one more caution is whether the machine you decide on has a history of problems. If so, you would want to make sure that the manufactuer still supports the machine. Another alternative may be to try to collaborate with someone at an institution that already has a sequencer. You could prepare all of you samples locally and then spend a bit of time in the lab of someone who has a bit of space and would be willing to sponsor you. Just another thing to consider. Good luck! Hi Andrea, I know of cheaper prices for sequencing work at Cardiff University - they are one of the cheapest services but still relatively expensive. There is also a sequencing service in Korea Macrogen but you have to ship it there and you really have to do above a certain number of runs before it becomes worth while but surprisingly if you have enough it can be one of the best options. In the UK there is a NERC facility that ytou can apply for a sheffield University but I'm not sure what the eligibility foir this facility is exactly. If you need further information on any of these let me know and I'll give you some more details. If you find out about any other good cheap facilities that are better than the ones I've listed please could you pass on the details. Good luck, Vittoria I am currently using a Li-cor system (4300) and I have previously used a Corbett Research GelScan2000. Both are very useable. The Gelscan is cheaper and I could push samples through quicker, but is only has one channel and, outside of Australia where Corbett is based, support is poor. The Li-cor is more expensive, may take longer (but could actually be as fast with the short plates. I only have the long plates) but it has two channels (i.e., you can run two different colored dyes at the same time) and probably has better support. The dyes are quite expensive for the Li-cor machine, whereas the Corbett machine uses the same dyes as PE machines. Hope that helps. Any further questions, drop me a line. Mike We use LiCor DNA sequencers to score microsatellites. They're reliable, and the customer support is outstanding (much better than ABI). www.licor.com Best wishes, Mohamed Noor --- evoldir@evol.biology.mcmaster.ca ha scritto: Hi everybody, Thankyou very much for your answers. I have attached a file containing them all. To summarize I found that not only I find it difficult and quite subjective to score AFLP peaks. (thanks that was nice to hear). The advise adds up to making the DNA quality as good as possible. Running at least some samples twice to check the error percent. Try to standazise the samples amplification. Choose a minor level eg. 10 percent of largest peak. And ofcause it is advisable to mix samples on gels and scoring without knowing the origin of the sample. Also I will score some more peaks so I have at least 125 good ones for my analysis. Thankyou again for your help. Marianne Lindhardt mariannel@bi.ku.dk PER FAVORE RISPONDETE A QUESTO EMAIL verardino@gmail.com PLEASE REPLY TO THIS EMAIL verardino@gmail.com POR FAVOR CONTESTAR A ESTE EMAIL verardino@gmail.com Dr Andrea Verardi Dipartimento di Genetica e Biologia Molecolare Universitą La Sapienza Roma mobile (++39) 340 97 17 555 email verardino@yahoo.com andrea verardi