I am PCR amplifying the ITS region of endophytic fungi using primers ITSF1/ITS4 and ITS1/ITS4-Tul. I know the amplicon length is highly variable, but I am trying to find a reasonable ballpark, at least for Tulasnella. I cannot find this information in the literature. Can anyone provide any assistance?
To find out:
1. Download all Tulasnella / Epulorhiza sequences from GenBank
2. Run ITSx (http://microbiology.se/software/itsx/) on them
The others are right when they say that (some) Tulasnella/Epulorhiza have truly divergent ITS regions.
It should be about 613 bp (estimated in silico using ITS1+4 on gb|GU166405.1).
Good luck.
Check publications by Martin Bidartondo and/or Lee Taylor - there should be some information in those.
cheers
The length of ITS sequence amplified by primers ITS 1 and ITS 4 ranged 600 to 750 bp.
I agree to the previous comments. Expect some 580-750bp if using ITS1F-ITS4. If using ITS4Tul, the products will be longer (cca 60 bp). Good luck, Lee
Thank you all for your help. I was finding much more variability from the literature.
Rasmus Kjoller, I have read quite a bit of Lee Taylor, but I was wanting some confirmation elsewhere. Thank you for your suggestion of Martin Bidartondo.
It is possible that the Birth-and-Death is driving the evolution of this species, thus causing the variability in your amplicons.
You should check carefully your aligment , because ITS-5.8 in Tulasnnella is very variable. Probably you can find inclusively length in the aligment about 870 bp without gaps for some Tulasnella spp.
In some studies you can find different groups for ITS-5.8 for Tulasnella due to the variability of this region, for instance, (Suárez et al. 2006; Shefferson et al. 2007; Taylor and McCormick 2008; Nontachaiyapoom et al. 2010; Cruz et al. 2011)
I agree with Dario Cruz... There are some Tulanella sp. with big insertions.
To find out:
1. Download all Tulasnella / Epulorhiza sequences from GenBank
2. Run ITSx (http://microbiology.se/software/itsx/) on them
The others are right when they say that (some) Tulasnella/Epulorhiza have truly divergent ITS regions.
Hi ,
What will be the length of ITS sequence amplified by primers ITS 5 and ITS 4 ? range
It is variable according the species in study. However the length range between 600 to 900 bp. Please checkArticle High-Coverage ITS Primers for the DNA-Based Identification o...
http://lutzonilab.org/nuclear-ribosomal-dna/
Hi
DeepHypha http://ocid.nacse.org/research/aftol/primers.php
The Bruns lab http://plantbio.berkeley.edu/~bruns/primers.html
The Hall lab http://protist.biology.washington.edu/bio2/people/bio.html?parecID=142
The Vilgalys lab http://www.biology.duke.edu/fungi/mycolab/primers.htm
AFTOL https://aftol.biology.duke.edu/pub/primers/viewPrimers
- Badges
- Science method