|
| Status |
Public on Jan 25, 2017 |
| Title |
AGO1IP-Mock-br2 |
| Sample type |
SRA |
| |
|
| Source name |
13-day-old seedlings, with AGO1-IP
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
genotype/variation: His-FLAG-AGO10 transgenic
ip: AGO1
rip antibody: anti-AGO1 (Agrisera, AS09 527)
treatment: Mock treatment included buffer only (no SDN)
biological replicate: 2
age: 13 days old
developmental stage: seedling
|
| Treatment protocol |
AGO1- and AGO10-RISC were immunoprecipitated (IP) from His-FLAG-AGO10 transgenic plants using anti-AGO1 (Agrisera) and anti-FLAG (Sigma) antibodies, respectively. Beads containing AGO1- or AGO10-IP were resuspended in reaction buffer (50 mM Tris, pH 8.0, 150 mM NaCl, 1 mM DTT, 2.5 mM MnCl2, 1mM ATP) and then incubated with SDN1 or SDN1-D283A catalytic mutant for one hour. Mock treatment included buffer only (no SDN). Beads were collected after incubation for RNA extraction.
|
| Growth protocol |
13-day-old seedlings grown on ½ Murashige and Skoog (MS) plates under 16h light/8h dark cycle
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted with TRIzol reagent (Life Technologies, USA, 15596018)
Libraries were made using the NEBNext Small RNA Library Prep kit (NEB, USA) according to manufacturer’s instructions. Briefly, 3' and 5' adapters were ligated sequentially to the small RNAs. Ligated small RNAs were converted to cDNA by reverse transcription followed by PCR amplification for 15 cycles. The barcoded libraries were sequenced on the Illumina NextSeq500.
|
| |
|
| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
Col-HFAGO10-AGO1-IP-Mock-br2
|
| Data processing |
Raw reads were processed by first trimming 3’ adapter sequence using custom Perl script. Reads without 3’ adapter sequence or less than 18 nucleotides after trimming were discarded from further analysis.
Trimmed reads were aligned against a custom database containing rRNA, tRNA, snoRNA, and snRNAs using Bowtie 0.12.8 to filter out rRNA reads.
Trimmed reads were collapsed into unique sequences with counts (tag count files) and processed as described in Zhao et al. Curr Biol 2012 for determining 3' end truncation and tailing.
Genome_build: TAIR10
Supplementary_files_format_and_content: tag count files contain raw reads (after adapter-trimming) collapsed into unique sequences with the corresponding read count in the library. This file is used to analyze the modifications of miRNA 3'ends as described in Zhao et al. Curr Biol 2012.
|
| |
|
| Submission date |
Dec 28, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Meera Nair |
| E-mail(s) |
meera.nair@ucr.edu
|
| Phone |
951-827-7734
|
| Organization name |
University of California, Riverside
|
| Department |
Biomedical Sciences
|
| Lab |
3135 Multidisciplinary Building
|
| Street address |
3401 Watkins Drive
|
| City |
Riverside |
| State/province |
CA |
| ZIP/Postal code |
92521 |
| Country |
USA |
| |
|
| Platform ID |
GPL19580 |
| Series (2) |
| GSE87351 |
ARGONAUTE10 promotes the degradation of miR165/6 through the SDN1 and SDN2 exonucleases in Arabidopsis [AGO10SDN] |
| GSE87355 |
ARGONAUTE10 promotes the degradation of miR165/6 through the SDN1 and SDN2 exonucleases in Arabidopsis |
|
| Relations |
| BioSample |
SAMN06189252 |
| SRA |
SRX2448645 |
