Genomic dna clean and concentrator kit

Leaf tissues were collected from 189 RILs and parents (Cocodrie and Dular) grown in the control environment. The genomic DNA was extracted using a modified CTAB method [79 (link)]. Extracted DNA was purified by the genomic DNA clean and concentrator kit (Zymo Research Corp. Irvine, CA, USA). The DNA concentration in each sample was estimated by Nanodrop ND-100 Spectrophotometer (Thermo Fisher Scientific, Wilmington, USA). The concentration of DNA was adjusted to 30–60 ng/µL for library construction. The protocol reported by Elshire et al. [80 (link)] was used for library preparation using the ApeKI restriction enzyme followed by single-end sequencing at the Genomic Diversity Facility of Cornell University.
The TASSEL 3 GBS pipeline was used to process the raw sequence data [81 (link)]. Raw sequences without barcode were removed using the TASSEL plugin and good barcode reads were aligned to the Nipponbare reference genome using Bowtie 2 [82 (link)]. SNP calling and filtering were done using the TASSEL pipeline. The SNPs with high genotyping error, low coverage, and high heterozygosity were purged. Duplicate SNPs were merged. The remaining SNPs were manually filtered. SNPs markers with more than 10% missing alleles and SNPs with monomorphic alleles between both parents were discarded. The heterozygous SNPs were considered as missing data.

DNA was extracted from approximately 50 mg of homogenised oak tissue, using the DNeasy Plant Mini kit (Qiagen) according to the manufacturer’s instructions. Quality and concentration of samples were determined using agarose gel electrophoresis and the Qubit dsDNA HS assay kit (Thermo Fisher) according to the manufacturer’s instructions. In order to enrich microbiome DNA, the host DNA was depleted from the sample using the NEBnext microbiome DNA enrichment kit (New England Biolabs) according to the manufacturer’s instructions. Subsequently, the DNA was purified and concentrated using the Genomic DNA Clean and Concentrator kit (Zymo Research) according to the manufacturer’s instructions and stored at − 20 °C.

Genomic DNA was extracted from clinical isolates using the CTAB (cetyltrimethylammonium bromide) method (32 (link)) and purified using the Genomic DNA Clean and Concentrator kit (Zymo Research, Irvine, CA, USA). Purified DNA was assessed for quality and quantity using Nano DropTM and QubitTM dsDNA assay kit method (ThermoFisher Scientific, Waltham, MA, USA). Sequencing libraries were prepared using the NEBNext Ultra DNA Library preparation kit. In brief, fragmented DNA was subjected to a series of enzymatic steps for repairing the ends and tailing with dA-tail followed by ligation of adapter sequences. Adapter ligated fragments were cleaned up using SPRI beads, and the clean fragments were indexed using limited cycle PCR to generate final libraries for paired-end sequencing on a HiSeq X 10 sequencer (Illumina, San Diego, Ca, USA).