This is my first blog post in years - primarily down to a second child who is now a toddler. And what better topic to return to than a mainstay of past content than NCBI BLAST? This time with a motivating example from my recent work, metabarcoding. This is term used for sequencing a diagnostic region of DNA using specific primers for a group of organisms of interest, and then matching that amplicon to a database of known species. Human interpretation of a BLAST search can generally put a good guess as the organism - weighing hits and annotated taxonomy (e.g. ignoring the odd suspicious uncultu ..read more

All my recent blog posts have been looking at issues raised by the recent letter Shah et al. (2018) "Misunderstood parameter of NCBI BLAST impacts the correctness of bioinformatics workflows", and the associated test case which they ought to have included with the letter itself. Thus far I have focussed on: "What BLAST's max-target-sequences doesn't do" (2015 blog post, with links to follow up posts in 2018), an issue Suaji Kumar and I called a scary BLAST+ bug to do with alignment limits, which the NCBI BLAST team viewed as expected behaviour of a heuristic setting. "BLAST tie breaking by d ..read more

My November blog posts discussing the BLAST+ tools behaviour with an alignment limit setting (see What BLAST's max-target-sequences doesn't do, and the links from it), touched on database order, which comes into play as a tie breaker. Well, how is the BLAST database order defined? It turns out to be the reverse of the FASTA file used with makeblastdb, or in other words: Last-in, First-out (LIFO). Making simple test casesThe idea here is to make a database full of almost identical sequences, by adding a tiny barcode to the end. For the nucleotide example, I've used a 3bp barcode using all the ..read more

This is the fourth in a series of blog posts seeking to throw light some of the claims about the BLAST+ tool recently published by Shah et al. (2018) "Misunderstood parameter of NCBI BLAST impacts the correctness of bioinformatics workflows". It was very frustrating that the letter did not provide a reproducible test case, but in reply to the first pair of posts (one and two, both on Friday 2 November 2018), lead author Nidhi Shah got in touch via the comments on Sunday 4 November, with the URL to a GitHub repository describing the Shah et al. (2018) test cas ..read more

This is the third in a series of blog posts seeking to throw light some of the claims about the BLAST+ tool recently published by Shah et al. (2018) "Misunderstood parameter of NCBI BLAST impacts the correctness of bioinformatics workflows". It was very frustrating that the letter did not provide a reproducible test case, but in reply to the first pair of posts (one and two, both on Friday 2 November 2018), lead author Nidhi Shah got in touch via the comments on Sunday 4 November, with the URL to a GitHub repository describing the Shah et al. (2018) test case. Thank you! Their test ..read more

This is the second in a series of blog posts seeking to throw light some of the claims about the BLAST+ tool recently published by Shah et al. (2018) "Misunderstood parameter of NCBI BLAST impacts the correctness of bioinformatics workflows". Since regrettably they did not provide a reproducible test case, my previous post began by introducing a minimal test case. This topic dates back to 2015, when Sujai Kumar reported as a scaryBLAST+ -max_target_seqs bug which as I wrote about ("What BLAST's max-target-sequences doesn't do"), the NCBI BLAST developers explained it as a poorly doc ..read more

Back in 2015, my blog post "What BLAST's max-target-sequences doesn't do" highlighted what we called a scary BLAST+ -max_target_seqs bug, found and reported by Sujai Kumar. The NCBI BLAST teams took the stance this was a feature not a bug (and as a heuristic search tool, this is an understandable view), but conceded it could be better documented. Sadly, I don't think there has been much if any clarification in the BLAST+ documentation about the settings limiting the number of alignments returned, and what else they control. The recent letter Shah et al. (2018) "Misunderstood parameter of ..read more

Another quick bug report blog post, this time NCBI Entrez's espell is currently broken returning: Couldn't resolve #PubmedSpellSrv, the address table is empty. Here's an example in full: $ curl -v https://eutils.ncbi.nlm.nih.gov/entrez/eutils/espell.fcgi?term=biopythooon *   Trying 165.112.7.20... * TCP_NODELAY set * Connected to eutils.ncbi.nlm.nih.gov (165.112.7.20) port 443 (#0) * TLS 1.2 connection using TLS_ECDHE_RSA_WITH_AES_256_GCM_SHA384 * Server certificate: *.ncbi.nlm.nih.gov * Server certificate: DigiCert SHA2 High Assurance Server CA * Server certificate: DigiCert High ..read more

I've not managed to blog much at all this year (parenthood), so here's a quick BLAST+ bug report from working on updating the Galaxy wrappers: I've found a reproducible segmentation fault in tblastn under both Mac and Linux when requesting HTML output. Sample FASTA data files four_human_proteins.fasta and rhodopsin_nucs.fasta are examples I use often as test cases. This example works perfectly on BLAST+ 2.6.0, here on Mac OS X $ ~/Downloads/Software/ncbi-blast-2.6.0+/bin/tblastn -query four_human_proteins.fasta -subject rhodopsin_nucs.fasta -evalue 1e-10 -out tblastn_fou ..read more

Last summer I applied to the  Mozilla Fellows for Science 2016 call. Congratulations to the four 2016 fellows, selected from an impressive 483 submissions for only the second year of this innovative program. I was delighted to be short listed and interviewed, but also slightly relieved not to have made the final cut. This is due to the timing of personal circumstances - I'm now a father, and despite the taking time off under the UK's Shared Parental Leave scheme, I'm currently trying to cut back work related activities. While preparing my application, I was ..read more