Will you stop using 454?

[kmcarr]

What applications do you need 1000bp or more for that can't be broken down into short uniquely barcoded amplicons tiled across regions?

Plant transcriptomics. With so many recent segmental or whole genome duplications in plants assembling paralogous transcripts from shorter reads can be maddening.

[lletourn]

16s metagenomics...instead of having individual regions, you'll get almost all regions in each reads. Great for measuring diversity, richness, etc.

Also longer reads really make a difference when assembling. Mate pairs make a bigger difference, but when combining both...

[james hadfield]

750-1000bp reads are very useful but with only 1-1.5M reads is it better sticking to 200-400bp and getting maybe 3-4M of them on MiSeq?

I do not run any long read projects, we did a couple of Metagenomics experiments where long reads were more helpful. However a groupjust published a comaprison of 454 to Ilumina for this and the increased number of reads seemed a pretty strong reason to use Ilumina. This did not take account of possible 200-400bp mini-contigs.

What applications do you need 1000bp or more for that can't be broken down into short uniquely barcoded amplicons tiled across regions?

None of these companies stand still.

That's what makes our jobs so much fun!

[avilella]

I wonder how long Roche 454 will still be around if Ion gets to 400bp. Also if Illumina's chemistry performs as well as it looks like it will then 200-300bp is not unreasonable, coupled to PE reads we could see Illumina at 400bp mini-contigs.

With two short-read platforms offering close to Roche 454 read-lengths but with higher numbers of reads, lower costs, and in Illumina's case the simplicity of sample prep, when will we stop using 454?

454 plans to launch a new sequencing chemistry for the GS FLX that will reach read lengths to 750 bases and beyond. Supposedly by the end of June...

[lletourn]

We've been using, as beta, their long reads and they work fairly well. I can't give numbers but we had reads that hit the 1kb mark.

Even at 700b the median was over Q20.

Roche is not standing still for sure. When Ion torrent hits the 400b, I'm gussing 454 will already be at the 1k, 1.2k mark.

[maubp]

But Roche won't stand still - I recall hearing something about 1kb read 454 upgrade being tested... does anyone know more?

[nickloman]

We'll probably keep using it for amplicons (particularly 16S) until a cheaper platform gives reasonably accurate long reads above 400bp. Ion Torrent are promising this sometime in 2012. Illumina may get there around the same time e.g. a 2 x 200bp paired read is not too difficult to imagine. This would take a long time on the HiSeq but perhaps not on the MiSeq.

[james hadfield]

I wonder how long Roche 454 will still be around if Ion gets to 400bp. Also if Illumina's chemistry performs as well as it looks like it will then 200-300bp is not unreasonable, coupled to PE reads we could see Illumina at 400bp mini-contigs.

With two short-read platforms offering close to Roche 454 read-lengths but with higher numbers of reads, lower costs, and in Illumina's case the simplicity of sample prep, when will we stop using 454?