tissue: mixed stage inflorescence fraction: total antibody: n/a
Treatment protocol
These samples are from immunopurifications of either AGO4, AGO6 and AGO9 in Arabidopsis thaliana or small RNAs isolated, cloned and sequenced from Arabidopsis.
Growth protocol
standard growth conditions (16hr light, 21 C)
Extracted molecule
total RNA
Extraction protocol
The extract protocol varies for total sRNA and for immunopurifications (IP). If IP, proteins were purified with appropriate antibody and associated RNA was purified with Trizol reagent. If total sRNA, total RNA was extracted using Trizol reagent and small RNAs purified using mirVana (Ambion). For both of these types of samples, a 5' adapter was ligated, gel purified, followed by 3' adapter ligation, gel purification, 1st strand synthesis of RNA to create a cDNA library which was amplified with approximately 15 cycles of PCR. The PCR amplified library was then gel purified.
Library strategy
RNA-Seq
Library source
transcriptomic
Library selection
size fractionation
Instrument model
Illumina Genome Analyzer II
Description
small RNA sequencing Total sRNA sample; biological replicate with SL25 and technical replicates SL23_1 and SL23_2 raw data file: SL24_1.fq (fastq file)
Data processing
Small RNA adapters were removed using an exact match to the first 8bp of the 3' adapter (or the barcode + 3' adapter for multiplexed samples SL234B/C/F and SL235B/H). After adapter removal, reads were aligned to the Arabidopsis thaliana genome (TAIR v8) using SSAHA v3.1.
