DOI: 10.5061/dryad.bcc2fqzmb
title: Effective seed sterilization methods require optimization across maize genotypes

description: Studies of plant-microbe interactions using synthetic microbial
 communities (SynComs) often require the removal of seed-associated
 microbes by seed sterilization before inoculation to provide gnotobiotic
 growth conditions. A diversity of seed sterilization protocols have been
 developed in the past and have been used on different plant species with
 various amounts of validation. From these studies, it has become clear
 that each plant species requires its own optimized sterilization protocol.
 It has, however, so far not been tested if the same protocol works equally
 well for different varieties and seed sources of one plant species. We
 evaluated six seed sterilization protocols on two different varieties
 (Sugar Bun & B73) of maize. All unsterilized maize seeds showed
 fungal growth upon germination on filter paper, highlighting the need for
 a sterilization protocol. A short sterilization protocol with hypochlorite
 and ethanol was sufficient to prevent fungal growth on Sugar Bun
 germinants, however, a longer protocol with heat treatment and germination
 in fungicide was needed to obtain clean B73 germinants. This difference
 may have arisen from the effect of either genotype or seed source. We then
 tested the protocol that performed best for B73 on three additional maize
 genotypes from four sources. Seed germination rates and fungal
 contamination levels varied widely by genotype and geographic source of
 seeds. Our study shows that consideration of both variety and seed source
 is important when optimizing sterilization protocols and highlights the
 importance of including seed source information in plant-microbe
 interaction studies that use sterilized seeds.
description: # Effective seed sterilization methods require optimization across maize
 genotypes
 [https://doi.org/10.5061/dryad.bcc2fqzmb](https://doi.org/10.5061/dryad.bcc2fqzmb) Excel files 'Experiment 1' and 'Experiment 2' contain raw data. R markdown file 'Seed_Sterlization_Manuscript' contains R code used for data analysis. Text files 'B73' and 'SugarBun' were used in the R script. #### Descriptions of data in Excel files 'Experiment 1' and 'Experiment 2': * The genotypes used in the study were ((all were used in experiment 2, only B73 and Sugar Bun were used in experiment 1):  1. Inbred varieties: B73, Mo17 2. Modern (hybrid) varieties: Sugar Bun (SB), Mo17 x B73, B73 x Mo17 * Several sources of seed were used for experiment 2. Each case would have a source as well as a genotype used (see above): 1. Clayton field seed harvested in 2021: CL21 2. Clayton field seed harvested in 2022: CL22 3. Seeds harvested from a greenhouse sown in 2021, harvested in 2022: GH21/22 4. Seeds harvested from Mexico fields (sown in 2021): MX21/22 * Germination and contamination were recorded for each seed in a 24-well plate. Positive for germination is recorded as "1", contamination is recorded as "1". The letters (A-D) and numbers (1-6) represent the coordinates for each seed in the plate.  * For experiment 1, two seed sterilization protocols were tested, a 'Long' and a 'Short' protocol. Variations in each protocol included a 'Pre Soak' step, and the germination of sterilized seeds on 'Fungicide'.