Unable to classify OTU with cluster.split

Good day to you Pat and Mothur team.

I have been encountering this error for a month and I’m stuck thus far. I’m trying to classify OTU’s in my 18 microbial marine biofilm samples but have always faced system failure (mouse froze, all programs stop, no response to commands) before I’m forced to soft restart the PC.

Most of the commands were following in accordance to MISEQ SOP with the exception of using the latest trainset data from Silva database.

I have used two different computers (pc and laptop), but both failed at the cluster.split step so far.
My computer specs are as follows:
|Processor|AMD Ryzen 5 2600 Six-Core Processor 3.85 GHz|
|Installed RAM|16.0 GB|
|System type|64-bit operating system, x64-based processor|

I have also reduced the number of samples to run Mothur (9 samples collectively were of a different location from the other 9) to reduce the data load, so it is possible to run them separately but no dice.

From the logfile it seems to be running fine until stopping abruptly, no error message whatsoever. Should I continue running Mothur beyond the amount of hours allocated? (It was close to 4 hours after cluster.split was initiated before initiating pc restart). For reference, there were close to 1692466 unique sequences.

The full logfile can be found here:

So, do I need to start looking for an even stronger PC or is there other ways to perform mothur without breaking the bank?

Hi - it looks like you used the V3 chemistry and/or are not sequencing the V4 region (can you clarify what you’re doing). Either of these will cause problems with data quality that lead to an inflated number of sequences like you are seeing. The result is processes like cluster.split crashing. I’d encourage you to check out this blog post…