We have designed a panel with 28 amplicons with a average length of 143. We already have 96 different barcodes from the IonXpress kit, but we can assure enough reads for each barcode/sample, so how many samples in this case, can be run at the same time in a 3.18 chip? Can we run the 96 selected samples (with the 96 barcodes) at the same time?
Yes, just bare in mind that the coverage can be low. In my experience the 318 chip is the one with the least bang for your buck, especially if your doing it in house since it requires 2 times the reagents of 316 and does not give the same amount of coverage (i think in practice it's less) you get from running 2 316 chips. Also unless i'm missing something I don't understand why use 96 bar codes on 28 samples. And one last thing, I have to say that I was really underwhelmed by the quality and quantity of the ion-torrent reads and really recommend you to switch to miseq if you have the option.
It depends on the depth of coverage that you need, but with the small number of amplicons that you are studying, I think you would be fine running all 96 samples on one 318 chip. There is a formula that you can get from tech support (maybe also from the Ion Community), for calculating the number of samples you can run per chip depending on the desired depth of coverage. I'm sorry that I don't have the formula handy. Nonetheless, we routinely run 6-8 samples per 318 chip for >66000 amplicons, and get ~3-7 miliion >=Q20 bases. We also run 2 whole exome libraries per 318 chip with good results. Although the uniformity of coverage, even within a batched run, is more variable than that of the MiSeq, we are mostly happy with the quality of the data from our Ion Proton platform, especially the data that we have generated using the new high sensitivity sequencing kit, which incorporates the a high fidelity polymerase. We have stuck with the Ion Torrent PGM/Prototon platform through all of the growing pains of the technology because of the scalability of the technology.
Yes, I agree with Julie, load all 96 samples on one 318 chip, and if you don't have enough coverage after all, just re-sequence the pool (on a 316 or 318, depending on how much more coverage you need). This way you also avoid batch effects from splitting the samples for different runs.
Thank you very much for your answers, they have been very helpful. We will try with 1 3.16 chip, because the coverage we get is more or less acceptable.
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