Cheapest solution for targeted sequencing?

sulicon · 08-12-2011, 02:21 PM

Hi,

We have 500-1000 genomic DNA samples and we are interested in a set of ~20 loci. What's the most affordable way to sequence the 20 loci for all the samples?

Thanks,
Shuli

ulz_peter · 08-15-2011, 11:58 PM

If it's only 20 Loci I would do PCR and Amplicon sequencing. (Well, depends on the size of the Loci). You could use multipliex identifiers of any length which you incorporate into the amplification primer. With that amount of samples you could also think about a multiplex PCR amplifying 10 or even 20 PCR products in one reaction: that would lead to one PCR per sample targetted sequencing: I don't think anyone can do it cheaper...

rocksd · 09-02-2011, 06:32 AM

Quote:

Originally Posted by sulicon

Hi,

We have 500-1000 genomic DNA samples and we are interested in a set of ~20 loci. What's the most affordable way to sequence the 20 loci for all the samples?

Thanks,
Shuli

How long (bp) are those loci? If it is short enough, I'd say perform multiplex PCR using primers embedded with nextgen adaptors. In this case, sample prep cost may be significantly lower. You can multiplex many samples to control sequencing cost (sequencing reagents and materials), depending on the size of the loci. But the library prep may be too much a burden if you follow Illumina's gDNA library protocol for 500-1000 samples. On the other hand, if you can generate enough amplicons with Adaptors attached already, it will be a lot less work.

I am dealing with a similar request from another client. If you wish to discuss a little more on this topic, you can send me a personal message.

Rocksd

stanford_genome_tech · 09-24-2011, 11:04 PM

Hi all,

Stanford Genome Technology has published a 2nd generation "open source" capture technology using in solution oligonucleotides.

We have an in silico design covering the CCDS genes (most of the exome) release and will have one covering the majority of the genome out soon (under review). You can download oligo sequences and create your own capture assays.

http://oligoexome.stanford.edu

http://www.plosone.org/article/info%...l.pone.0021088

More completely new third generation targeting technologies coming soon in a Nature journal. Be on the look out for it shortly.

SGTC sequencing techies

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08-12-2011, 02:21 PM	#1
sulicon Member Location: Los Angeles Join Date: Aug 2010 Posts: 41	Cheapest solution for targeted sequencing? Hi, We have 500-1000 genomic DNA samples and we are interested in a set of ~20 loci. What's the most affordable way to sequence the 20 loci for all the samples? Thanks, Shuli

08-15-2011, 11:58 PM	#2
ulz_peter Senior Member Location: Graz, Austria Join Date: Feb 2010 Posts: 219	If it's only 20 Loci I would do PCR and Amplicon sequencing. (Well, depends on the size of the Loci). You could use multipliex identifiers of any length which you incorporate into the amplification primer. With that amount of samples you could also think about a multiplex PCR amplifying 10 or even 20 PCR products in one reaction: that would lead to one PCR per sample targetted sequencing: I don't think anyone can do it cheaper...

09-24-2011, 11:04 PM	#4
stanford_genome_tech Junior Member Location: northern california Join Date: Aug 2009 Posts: 7	Hi all, Stanford Genome Technology has published a 2nd generation "open source" capture technology using in solution oligonucleotides. We have an in silico design covering the CCDS genes (most of the exome) release and will have one covering the majority of the genome out soon (under review). You can download oligo sequences and create your own capture assays. http://oligoexome.stanford.edu http://www.plosone.org/article/info%...l.pone.0021088 More completely new third generation targeting technologies coming soon in a Nature journal. Be on the look out for it shortly. SGTC sequencing techies