Poster: Biology & Disease Mgmt: Nematology
Comparison of ITS1 versus ITS2 regions for characterizing fungal communities colonizing soybean cyst nematode
W. HU (1), K. Bushley (1), S. Chen (2) (1) Department of Plant Biology, University of Minnesota, U.S.A.; (2) Southern Research and Outreach Center, University of Minnesota, U.S.A.
The development of next generation sequencing has enabled investigation of microbial communities from a wide range of environmental samples. We examined fungal communities associated with cysts of soybean cyst nematode (SCN) to understand the effects of different crop rotation practices. The primers most frequently used for fungal communities on the Illumina Miseq platform target either the rRNA internal transcribed spacer 1 region (ITS1) or the ITS2 region. In order to assess which primers are better for amplifying fungal communities in SCN cysts, we performed 300 bp paired-end sequencing on two Miseq runs using fungal ITS1f/ITS2r and 5.8SR/ITS4 primers. Cysts of the SCN were collected from a soybean-corn rotation field in Waseca, MN, and washed with 0.2% Tween™ 20. The DNA was extracted from 50 cysts using the CTAB method, and normalized for PCR reactions. After quality control, 361224 (ITS1) and 230404 (ITS2) reads were used for OTU picking in QIIME. OTUs were blasted against the UNITE database with the default setting in QIIME. About 99% of the ITS1 region sequences were classified as fungal sequences, however, 99% of the ITS2 region sequences were not fungal sequences. In addition, significant changes in the relative abundance of some taxa was observed between rotation treatments for only the ITS1 sequences. We conclude that primers targeting the fungal ITS1 region rather than ITS2 region are more suitable for characterizing fungal communities in SCN cysts.