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Sample GSM5087717 Query DataSets for GSM5087717
Status Public on Nov 23, 2021
Title ddm1-2_2C_SmartSeq2_S7_10B
Sample type SRA
 
Source name cotyledons
Organism Arabidopsis thaliana
Characteristics dapi fluorescence peak: 0B
developmental stage: two-week-old
tissue: cotyledons
genotype: ddm1-2
cell compartment: nuclei
Extracted molecule polyA RNA
Extraction protocol Fresh nuclei isolated from two-week-old cotyledons grown on 1%MS under long day conditions were isolated as follows. Roughly 50 cotyledons were finely chopped with razor into 50 ul Partec nuclei extraction buffer (Sysmex America, 05-5002), and stained with 400 ul Partec nuclei staining buffer. Samples were filtered once through a 35um nylon mesh (Falcon, 352235) and subjected to fluorescent activated nuclei sorting. Nuclei were sorted from 2C, 4C, 8C and 16C peaks based on DAPI fluorescence. 50 nuclei from each peak were sorted into individual wells of a 96 well plate. Three to four replicates per ploidy/genotype were collected. Negative controls, with no nuclei sorted into a well, were included in every plate and carried through library preparation and sequencing.
Libraries were prepared according to Smart-seq V2 protocol at reduced volume and with few modifications as described previously (Picard, C,L et al, bioRxive, 2020).
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NovaSeq 6000
 
Description S7_10B
Data processing Reads were aligned to TAIR10 with chloroplast and mitochondria excluded using hisat2 with default settings. Samtools view was used to filter uniquely mapped proper pairs with the following parameters: -b -q 60 -f 2. Duplicates reads were removed using Picard tools MarkDuplicates.jar (https://broadinstitute.github.io/picard/). Counts for each gene were generated using htseq-count using Araport 11 genes and TEs annotations. FPKM values and differential gene expression were analyzed using Cuffdiff with default settings. CummeRbound package was used to generate a boxplot of FPKM across sample replicates, a dendogram of Jensen-Shannon distance between sample replicates and the multi-dimensional scaling (MDS) plot for sample replicates for all genes. Poor quality libraries were filtered out, 2-4 replicates per ploidy/sample were retinaed and used in analysis.
Genome_build: TAIR10
Supplementary_files_format_and_content: Text file includes counts for each gene generated by htseq-count.
 
Submission date Feb 16, 2021
Last update date Nov 23, 2021
Contact name magdalena ewelina potok
Organization name UCLA
Department MCDB
Lab Jacobsen lab
Street address 4045 Terasaki Life Sciences Building
City Los Angeles
State/province CA
ZIP/Postal code 90095-7239
Country USA
 
Platform ID GPL26208
Series (2)
GSE166894 The role of ATXR6 expression in modulating genome stability and transposable element repression in Arabidopsis [SMARTSeq2]
GSE166897 The role of ATXR6 expression in modulating genome stability and transposable element repression in Arabidopsis
Relations
BioSample SAMN17927660
SRA SRX10112970

Supplementary file Size Download File type/resource
GSM5087717_S7_10B_unique_pairs_dedup_count_TEs.txt.gz 56.0 Kb (ftp)(http) TXT
GSM5087717_S7_10B_unique_pairs_dedup_count_genes.txt.gz 110.5 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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