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| Status |
Public on Dec 31, 2019 |
| Title |
WT_19 [small RNA] |
| Sample type |
SRA |
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| Source name |
wild type_pollen
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| Organism |
Arabidopsis thaliana |
| Characteristics |
accession: Col
genotype/variation: wild type
tissue: pollen
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| Extracted molecule |
total RNA |
| Extraction protocol |
Following the sorting of pollen grains, whole RNA was isolated using the Direct-Zol™ RNA MiniPrep Kit by Zymo Research (Irvine, California, US) since the specific columns provided in this kit prevent loss of small RNAs. The freshly sorted samples were centrifuged for 3 min at 3,000 rpm and the supernatant removed. Subsequently, the pollen were resuspended in 750 μl Tri-reagent LS (Sigma-Aldrich, St. Louis, US) and flash frozen in liquid nitrogen. The samples were ground in a cooled mortar to break up pollen grains until the samples liquefied and transferred to fresh reaction tubes. Afterwards, the protocol was followed according to the manufacturer’s specification. RNA concentration was measured using a NanoDrop and analysed on a BioAnalyzer to verify the existence of small RNAs as well as to check for possible RNA degradation.
Generation of small RNA libraries from three paired pollen RNA isolations (wild-type and paps1-3 mutant pollen sorted from the same sample) and sequencing was carried out by Eurofins Medigenomix GmbH, Ebersburg, Germany. Complementary DNA (cDNA) libraries were constructed containing fragments with a length of approximately 19-25 bp with TruSeqTM Small RNA Sample Preparation Kit (Illumina, San Diego, US) and sequencing was performed using Illumina HiSeq 2500.
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| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
Sample19_CAGATC_L005
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| Data processing |
RA3 adapters were removed using Trimmomatic (Bolger et al. 2014; doi:10.1093/bioinformatics/btu170).
The resulting reads were mapped against the Arabidopsis thaliana reference genome (TAIR10) using bwa (Li et al. 2009; doi:10.1093/bioinformatics/btp324) and further processed using samtools (Li et al. 2009; doi:10.1093/bioinformatics/btp352). Bam files were split by read length for 18 to 28 nucleotides using reformat.sh (https://sourceforge.net/projects/bbmap/).
Reads mapping to TAIR10 annotated gene exons and transposable elements (excluding overlaps with the prior ones) were counted using bedtools multicov (Quinlan and Hall 2010; doi:10.1093/bioinformatics/btq033).
Genome_build: TAIR10 (www.arabidopsis.org)
Supplementary_files_format_and_content: Raw read counts by length (18 to 28)
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| Submission date |
Mar 07, 2019 |
| Last update date |
Jan 01, 2020 |
| Contact name |
Michael Lenhard |
| E-mail(s) |
michael.lenhard@uni-potsdam.de
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| Phone |
+49-331-9775580
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| Organization name |
Universität Potsdam
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| Department |
Institut für Biochemie und Biologie
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| Street address |
Karl-Liebknecht-Str. 24-25, Haus 26
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| City |
Potsdam |
| ZIP/Postal code |
14476 |
| Country |
Germany |
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| Platform ID |
GPL17639 |
| Series (2) |
| GSE127992 |
The poly(A) polymerase PAPS1 interacts with the RNA-directed DNA methylation pathway in sporophyte and pollen development [small RNA] |
| GSE127993 |
The poly(A) polymerase PAPS1 interacts with the RNA-directed DNA methylation pathway in sporophyte and pollen development |
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| Relations |
| BioSample |
SAMN11082178 |
| SRA |
SRX5491754 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3659806_Sample19-counts.txt.gz |
609.9 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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