[GW_OK]

Well, as Genomax has posted from the user guide, the first 2 bases are the critical cluster calling cycles. I'm going to go find some data and count the number of reads with GG in the start. I will wager that it's a low percentage.

But, to paraphrase Gordon Gecko: Cynicism is good.

[jwfoley]

The reps in the webinar just said a non-G is required in the first five bases of the read, but for simple statistical reasons that's not likely to be much of a problem.

However, I worry about mild bias against reads that are G-rich in their whole length, not just severe bias against ones that are G-rich at the beginning. This could be particularly troublesome for specific applications like CpG methylation profiling.

[GW_OK]

Dear Illumina,
Please release a high-GC content data set from the NextSeq to BaseSpace.

Thanks,
SeqAnswers readership

[pmiguel]

Well, as Genomax has posted from the user guide, the first 2 bases are the critical cluster calling cycles. I'm going to go find some data and count the number of reads with GG in the start. I will wager that it's a low percentage.

But, to paraphrase Gordon Gecko: Cynicism is good.

What? Does the NextGen do clustering on the index read? (The SOLiD actually did. EDIT: wait, it did the index read first, but it identified clusters (beads actually) using a scan from a fluor annealed to all the beads) If not, then the task is much simpler -- just find a previously identified cluster and call bases. The fact that the NextGen can't handle 2 initial G's in the index read, suggests it is going to be much less tolerant of G's during clustering.

--
Phillip

[ericchow]

According to the NextSeq spec sheet (http://res.illumina.com/documents/pr...nextseq500.pdf), in addition to the 520 and 650nm LEDs, it also has a 780nm laser. At first I thought this might be for the reagent scanner, but the MiSeq spec sheet doesn't list it and the HiSeq spec sheet lists the scanner as 650nm.

Could this infrared laser be used to detect all clusters?

[pmiguel]

Anyone want to comment on the differences between a single HiSeq X Ten and a HiSeq 2500 1T?

There isn't much point trying to run a 1T lab -- same type of data for 4x the price. Unless it is just a matter of time before 1Ts will be running X Ten chemistry.

--
Phillip

[GenoMax]

Following is pure speculation so take it with a grain of salt.

I expect to see patterned flowcells trickle down to the 2500's (and 1500?) at some point in time this year (this is the upgrade most of us are waiting for) though it remains to be seen if a "new camera requirement" will go with it, which could leave a number of machines out in the cold. It is difficult to tell (from pictures) if the patterned flow cells are the same dimension as the current flowcells. If they are not then that could be yet another problem (unless illumina makes a new version of patterned flowcells for existing machines).

We know that 1T machines will use a new version of SBS reagents (v.4) along with some new software/firmware. Is that kit anything like what the X will use, anyone?

[TonyBrooks]

Following is pure speculation so take it with a grain of salt.

I expect to see patterned flowcells trickle down to the 2500's (and 1500?) at some point in time this year (this is the upgrade most of us are waiting for) though it remains to be seen if a "new camera requirement" will go with it, which could leave a number of machines out in the cold. It is difficult to tell (from pictures) if the patterned flow cells are the same dimension as the current flowcells. If they are not then that could be yet another problem (unless illumina makes a new version of patterned flowcells for existing machines).

We know that 1T machines will use a new version of SBS reagents (v.4) along with some new software/hardware. Is that kit anything like what the X will use, anyone?

http://www.illumina.com/systems/hiseq_2500_1500/performance_specifications.ilmn

v4 is mentioned here for the 1500/2500's. Looks like a higher cluster density and a corresponding bump in output (up to 2 billion reads per flowcell).

[GW_OK]

http://www.illumina.com/systems/hise...fications.ilmn

v4 is mentioned here for the 1500/2500's. Looks like a higher cluster density and a corresponding bump in output (up to 2 billion reads per flowcell).

From the fine print on the SBS v4 page:

*Compatible with factory-built HiSeq 2500 and 1500 units and select HiSeq 2000 units. Please contact Customer Service to confirm system compatibility.

So v4 is Hiseq1T chemistry only.

[pmiguel]

Following is pure speculation so take it with a grain of salt.

I expect to see patterned flowcells trickle down to the 2500's (and 1500?) at some point in time this year (this is the upgrade most of us are waiting for) though it remains to be seen if a "new camera requirement" will go with it, which could leave a number of machines out in the cold. It is difficult to tell (from pictures) if the patterned flow cells are the same dimension as the current flowcells. If they are not then that could be yet another problem (unless illumina makes a new version of patterned flowcells for existing machines).

We know that 1T machines will use a new version of SBS reagents (v.4) along with some new software/firmware. Is that kit anything like what the X will use, anyone?

Yes, that is the question: does SBS v4 + patterned flowcell cluster kit = running on a HiSeq "1T" == a HiSeq X Ten single instrument?

v4 (if your instrument can run it) gets you to $28/billion bases (reagent costs). Does the patterned flowcell take you to $7/billion bases (reagent costs)?

In other words, say I am a late Cretaceous dinosaur watching a light in the sky heading for Chicxulub. Am I avian, or non-avian?

--
Phillip

[GenoMax]

Yes, that is the question: does SBS v4 + patterned flowcell cluster kit = running on a HiSeq "1T" == a HiSeq X Ten single instrument?

We are assuming that there will be a "patterned" flowcell kit for 2500. I found this blog post (skip to the "Edit" at the bottom of the post), which says that "patterned" flowcells will only work with HiSeq X and TruSeq Nano kits.

Let us hope that is not true.

[pmiguel]

We are assuming that there will be a "patterned" flowcell kit for 2500. I found this blog post (skip to the "Edit" at the bottom of the post), which says that "patterned" flowcells will only work with HiSeq X and TruSeq Nano kits.

Let us hope that is not true.

The patterned flowcell may only work with the HiSeq X Ten, but I fail to see how a TruSeq Nano library molecule is distinguishable from any other (modern) Illumina library molecule.

--
Phillip

[austinso]

We are assuming that there will be a "patterned" flowcell kit for 2500. I found this blog post (skip to the "Edit" at the bottom of the post), which says that "patterned" flowcells will only work with HiSeq X and TruSeq Nano kits.

Let us hope that is not true.

Based on the flow-cell description for the HiSeqXTen, my guess is that significant mods would be required to allow 1. current to be linked the electrodes of the flow cell and 2. to control the current and the fluidics in a way to permit ordered cluster generation...

FWIW

[pmiguel]

My Illumina sales rep told me:

(1) Patterned flowcells are only for the HiSeqX.
(2) As previously reported only HiSeqs with certain serial numbers can run the v4 SBS chemistry. There is no upgrade path for HiSeqs outside that serial number range.
(3) The HiSeq X Ten may only be used for sequencing human genomic DNA libraries constructed with TruSeq Nano DNA prep kits. Not sure how Illumina can enforce this, but the word is that it will.

--
Phillip

[GenoMax]

Those who want/could use patterned flowcells need to start hoarding (thousands of) dollars. That way one will be ready when HiSeq X becomes available for purchase as a stand-alone instrument.

In case you had missed this from yesterday: http://seqanswers.com/forums/showthread.php?t=40314