I am looking forward to analyze changes in soil bacterial community by 16S amplicon sequencing.
Previously there were more reports on targeting V1-V2 region with primer sets of 27F/514R(or something similar). But recent reports e.g. Klindworth et al 2012 and Thijis et al 2017 suggest usage of V3-V4 region preferably with primer sets 341F/785R(or 805R). Is here any specific reason for change in targeting regions?
Reports suggest this region(primer set) gives better coverage and high phyla spectra but somehow the aforementioned primer set wasn't included in the comparative studies.
May be i am missing something. All help will be appreciated.
Hi
As you said some evidences from Klindworth et al 2012 that the better in some several aspects comparing to using other variable regions. It might be not completely true when you go with the sequencing large numbers of samples (you can check the http://blog.mothur.org/2014/09/11/Why-such-a-large-distance-matrix/)
Another thing, it is something to do with the miSeq platform from Illumina, we also were adviced to use the V3-V4 region for our analyses, but at the end of the day, we had many things to deal with, then we recognize that using single variable region such as V3 and V4 could be a better option
The shifts in the popularity of the various 16S hypervariable regions over time has a technical reason but also a historic reason to it, sort of like how the pant cuts have changed over the years in the fashion business or the shape and styles of the car headlights. The V4 region seemed to be very popular in the qiime community, and given that lots of people use qiime, lots of people seemed to target the V4. Then came Miseq that could target two V regions, so V3-V4 became popular. Why not V4-V5? I wonder too, although V5 is shorter than V3 so I'm guessing that V3 could provide better taxonomic resolution than V5? I've had good results working with V5 and there are very highly conserved priming regions flanking V5. Marine microbiologists have traditionally targeted the V6 region and those whereabouts, while insect microbiologists often target the V1-2 region (if my memory is correct).
- Badges
- Science method