Why does Illumina have the extra +1 cycle?
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3.3 years ago
Michael ▴ 270

What is the reason for the "+1 cycle" for Illumina sequencing?

For ex. when you do 2x150bp the device will do 2x151 cycles. For 2x300bp it will do two times 301 cycles, and so on...

What is the reason for this?

NGS Illumina • 3.5k views
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3.3 years ago
GenoMax 144k

Since the last base lacks phasing information some sequencing facilities choose to do an extra cycle of sequencing.

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hmm... I do not get exactly how the last base lacks phasing information. EDIT: do you mean to estimate how many strands of the cluster are out of phase? -> hence estimating the quality of the basecall?

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Since the last base has nothing in front of it to compare (doh!) there is no phasing information available. This is an older blog post that describes phasing. As chemistry and software has improved over time this has become much less of an issue.

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What confusing here that this seems like an inner implementation detail. If the setup is 150 bp, why do we end up with 151. Why doesn't the facility do 149 cycles + 1 to produce 150bp as originally set up? Plus should one keep this last base? I can see how this is confusing.

I suspect that this extra base used to be unreliable and cut off. Is it possible that in the past you usually you would get 150bp even after the +1.

But nowadays that last base may be good, and is being kept and like fruit vendor in the market, topping off you basket with a little extra, the facility is also giving you a little more than you asked.

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Thanks much! I still do not understand the phasing argument completely, but never mind :)

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Operators set up the sequencing run to be for n cycles. Sequencer does not do n+1 by default. Some providers used to do n+1 cycles (perhaps still do) to address possible phasing/quality problem with last base. It is dangerous to push the sequencer to do n+1 on long PE reads, especially if you are doing 2D indexes. Sequencer may run out of reagents and that is not a pretty thing.

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