Transposable Elements Contribute to Activation of Maize Genes in Response to Abiotic Stress 

Genbank Project: PRJNA369205

Oka et al 2015, Nathan Stam lab

https://genomebiology.biomedcentral.com/articles/10.1186/s13059-017-1273-4

Method:

Tissue preparation:

The seed stock of the maize B73 inbred line used in this study was obtained from J. Gardiner (University of Arizona, Tucson, AZ, USA) in 2013. It was obtained from the North Central Regional Plant Introduction Station in Ames, IA, USA (order no.: 169545, accession: PI550473, lot: 94ncai02). It is from the same accession (PI 550473) that was used for the maize B73 genome sequencing project [41], but a different lot number because it was requested several years later. The B-I plant stock used in this study (W23) was obtained from V.L. Chandler (University of Arizona, Tucson, AZ, USA). Maize plants were grown in the greenhouse at two different locations: The Max Planck Institute for Plant Breeding Research in Cologne (MPIPZ) and the University of Amsterdam (UvA). At the MPIPZ, maize plants were grown for DNase-seq and RNA-seq. At the UvA, maize plants were grown for H3K9ac ChIP-seq and RNA-seq. At both locations plants were grown in soil under 16-h/8-h light/dark cycles at an average temperature of 23 °C. The plants were harvested at the V2 stage (two collars visible; V2-IST), V5 stage (five collars visible; V5-IST) or when the silks started emerging from the husks. The two tissues used for the RNA-seq, DNase-seq and ChIP-seq experiments were the inner stem tissue of V2 seedlings, which is composed of the seedling stem with the outer leaves and all exposed leaf blades removed, and the soft inner husk leaves surrounding the ear; the tough outer husk leaves were discarded.

Read mapping:

Because RNA-seq reads were inadvertently sequenced from the reverse strand, the reverse complement of the fastq reads were generated using the fastx tool fastx_reverse_complement with the command fastx_reverse_complement -z -v -i INFILE -o OUTFILE on uncompressed fastq files.

The reverse-complemented reads were mapped using STAR 2.5.3. Paremeters for these reads were:

STAR --outSAMattributes All --outSAMmapqUnique 10 --outFilterMismatchNmax 0 --outBAMsortingThreadN 4 --limitBAMsortRAM 5594394835 --outSAMtype BAM SortedByCoordinate

Bigwig files were generated using bedtools genomecov -split -scale 1.0 followed by kentUtils bedGraphToBigWig.