Hello,
I am facing a problem with 16s rRNA (7 hypervariable regions) sequence data analysis. I analyzed the data (fastq files) with Qiime2 using ASV approaches but finally didn’t identified species levels. Mostly it says uncutured bacteria. I used updated Silva database to compare. Whereas, using the same data I received species level identification from the ion reporter (Thermofisher Scientific). They said they used Green gene database. And their approach was OTU, not ASV. Why the same data did show different results?
Akthough this is well known that green gene database is not updated regularly and OTU approach is a back dated method, my question is why I was unable to identify at species level using Silva database and ASV approach?
Please help me with your expert advice and discussion. Thanks!
I think there are a couple things going on. My suspecion is that Thermo-Fisher has adapted the Greengenes database and done some custom classification to improve resolution. (Against the license, but ? Where as Silva has fewer classified sequences because it's a more general database. To be honest, I'd want to know more about how Thermo-Fisher developed their database, but I suspect that's proprietary. You may want to look at the previous thread about the Thermo-Fisher sequencing, if you haven't checked it out yet:
The brilliant @Nicholas_Bokulich has also suggested checking your classifier regions! What classifier have you used? Is it V4 specific? Full 16s? Some other classifier?
? Where as Silva has fewer classified sequences because it's a more general database. To be honest, I'd want to know more about how Thermo-Fisher developed their database, but I suspect that's proprietary. You may want to look at the previous thread about the Thermo-Fisher sequencing, if you haven't checked it out yet: