WT2N41_ATGCTAGAGA
tick metagenome
host
Ixodes dentatus
isolation_source
eastern deciduous forests usually dominated by oak and hickory with variable understory vegetation frequently including or dominated by invasive shrubs
collection_date
2013-06-28
geo_loc_name
USA:Illinois,DuPage,Waterfall Glen Forest Preserve
lat_lon
41.86726 N 88.14108 W
ref_biomaterial
primary publication
samp_collect_device
Ticks were collected by dragging using a 1 meter square cloth and stored in 70% ethanol.
samp_mat_process
A Individual ticks were rinsed in 70% ethanol, air dried, and homogenized using a TissueLyzer II (Qiagen, Germantown, MD) in 2 ml deep-well 96-well plates at 25 Hz for 2 x 3 minutes. Each well contained a 5 ml steel bead (Qiagen) and 50 μl genomic digestion buffer. After tick homogenization in 80 µl (nymphs) or 100 µl (adults) 5% chelex, samples were incubated 20 min at 56 degrees Celsius, followed by an eight minute boil, and centrifugation at 4000xg for 7 minutes. Multiplex PCR followed by RLB was then performed as in Fredericks et al. (2016). Due to the requirement of purified DNA for Fluidigm analysis, a standard phenol-chloroform extraction with subsequent ethanol precipitation was done. DNA samples were quantified on a Qubit 2.0 (Life Technologies) using the Qubit dsDNA HS Assay Kit. Gene target regions were amplified using Fluidigm and sequenced using Illumina® HiSeq Rapid sequencing kit version 2 on a HiSeq 2500 sequencer (Illumina®, San Diego, CA, USA) at the W. M. Keck Center for Comparative and Functional Genomics at the University of Illinois at Urbana-Champaign as described previously (Muturi et al., 2016; doi:10.1186/s13071-016-1299-6). Briefly, sample DNA was amplified on the Fluidigm® microfluidics PCR platform with appropriate linkers and sample barcodes. The final Fluidigm® products were pooled in equimolar ratio for sequencing and spiked with 10% non-indexed PhiX control library (Illumina®) and sequenced by 2 x 250 nt paired-end sequencing on the Illumina® HiSeq® Rapid system. Fastq files were generated with the bcl2fastq v2.17.1.14 Conversion Software (Illumina®). Finally, the raw reads were demultiplexed by primer and by sample with the Fluidigm® demultiplexing pipeline v 2.0.
source_material_id
ATGCTAGAGA
host_life_stage
nymph
host_tissue_sampled
whole body
BioSampleModel
Metagenome or environmental