Genome analyzer iix platform

Worms for genome extraction were cultured in NGM under well-fed conditions at 20 °C. High quality genomic DNA was extracted using a Gentra Puregene Tissue Kit (QIAGEN, Hilden, Germany) according to the manufacturer’s instructions. The genome sequences of KR314, RC301, CB4854, and N2 were determined on an Illumina Genome Analyzer IIx platform using an Illumina TruSeq DNA Sample Prep Kit (Illumina, San Diego, CA, USA), which generated from 100 to 140 million reads; each read length was 110 bp. The genome sequences of AB1, CB4852, CB4853 were determined on an Illumina Hiseq 2500 platform using a TruSeq DNA PCR-Free LT Sample Prep Kit (Illumina), which generated from 50 to 130 million reads; each read length was 150 bp. Over 90 % of the genome was sequenced at a 10X coverage, except for CB4853 in which about 30 % was sequenced at 10X coverage. The nucleotide sequences reported in this paper have been submitted to the DDBJ Sequence Read Archive under accession numbers DRA002599 and DRA004249.

Supplementary appendix S1, Supplementary Material online, lists the cloned strains analyzed in this study, which include 19 Tbb, 13 Tbr, 2 Tbg1, 1 Tbg2, and 4 Te strains, together with country of origin and host. DNA was extracted from cryopreserved isolates from the Swiss Tropical and Public Health Institute, Basel or the University of Bristol. All strains were isolated in previous studies in adherence with national and institutional guidelines and extractions carried out using either a Qiagen micro DNA kit (Qiagen Pty Ltd) or using standard phenol–chloroform protocols dependent on sample quality. Fragmentation and library preparation and sequencing (2 × 75 bp) were conducted at the Yale Center for Genome Analysis, using either an Illumina Genome Analyzer IIx platform (STIB809) or an Illumina HiSeq 2000 platform (all other isolates) (data will be submitted to the National Center for Biotechnology Information short sequence read database upon acceptance). Quality control of reads was conducted using FastQC (Andrews 2010 ).

Total RNA was extracted from young fresh leaves and different stage of flowers using RNeasy Plant Mini Kit (Qiagen) following by the RNA purification by RNeasy MiniElute Cleanup Kit (Qiagen), according to the manufacture's protocol. Equal amounts of RNA from each sample were mixed together for the subsequent steps of our experiments. For mRNA library construction and deep sequencing, at least 20 µg of total RNA samples were prepared by using the TruSeq RNA Sample Preparation Kit (Illumina) for Illumina sequencing on Genome Analyzer IIx platform at CapitalBio Corporation (Beijing, China). The high quality reads obtained in this study have been deposited in the NCBI SRA database (accession number: SRA111764).

cDNA libraries were constructed from 18 leaf and two floret samples using a TruSeq RNA Sample Preparation Kit (Illumina Inc., San Diego, CA, USA) following the manufacturer’s instructions. Library quality was confirmed using the Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA, USA) and quantified via quantitative real-time PCR (qPCR) (Illumina protocol SY-930-10-10). Clustering was performed using a TruSeq Paired-End Cluster Kit on a cBot (Illumina Inc.). Paired-end sequencing was performed on an Illumina Genome Analyzer IIx Platform with TruSeq SBS 36-Cycle kits (Illumina Inc.) following the manufacturer’s specifications. RNA-seq was performed with floret libraries in two separate lanes without biological replicates. The remaining 18 leaf libraries from six different accessions (Table 1), each with three biological replicates, were distributed randomly in the flow cell with three libraries per lane.
The raw data was converted to FastQ files containing 72-bp reads. Quality control was performed using the NGS QC Toolkit v2.3.3 [52 (link)]. Initially, high-quality reads (Phred quality score ≥ 20 in at least 75% of bases) and reads with more than 60 bases were selected. Subsequently, reads were trimmed at the 3′ end for the removal of barcodes.

WES data from 9 paired primary triple negative breast cancer (TNBC) samples [31 (link)] is used in this study. Reads are sequenced to approximately 30× coverage on Illumina Genome Analyzer IIx platform and mapped to the reference genome NCBI36/hg18 using BWA [32 (link)]. We download the data from European Genome-Phenome Archive (EGA) under accession number EGAS00001000132.