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| Status |
Public on May 22, 2024 |
| Title |
shRNAseq in W22 roots Rep2 |
| Sample type |
SRA |
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| Source name |
root tips
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| Organism |
Zea mays |
| Characteristics |
tissue: root tips
inbred line: W22
genotype: WT
developmental stage: 5 DAG
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| Growth protocol |
For collecting immature ears, W22 maize inbreds were grown in the CSHL Uplands Farm field in the summer until they reached the appropriate stage. The plants were collected from the field, and 5-10mm primary and secondary ear primordia were dissected in the lab then frozen in LN2 and stored at -80°C. For pollen samples, shedding tassels of field-grown plants as described above were bagged in the evening and mature pollen was collected the following day. After passing through a sieve to remove anthers, pollen was frozen in LN2 and stored at -80°C. For endosperm samples, ears of field-grown plants were sib-pollinated and collected 15 DAP. Endosperm was dissected in the lab, frozen in LN2 and stored in -80°C. For root tips samples, seeds were germinated on wet paper towels in a Pyrex dish kept in the incubator at 26°C in continuous darkness. After 5 days, 1-3 mm root tips were cut off with a razor blade on ice, frozen in LN2 and stored at -80°C. For coleoptilar nodes samples, seeds were germinated in flats in a long day (8h dark/16h light) growth chamber, 27°C day and 24°C night, and light at 130 μmoles at the top flat. After 5 days, seedlings were unearthed and 5 mm sections around coleoptilar nodes were dissected on ice, frozen in LN2 and stored at -80°C.
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| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted with Direct-zol RNA Miniprep Kit (Zymo Reserach).
RNAseq libraries were constructed from 1ug of total RNA with TruSeq Stranded Total RNA LT Kit, including RiboZero Plant depletion and oligo(dT) and random probes for cDNA synthesis. RAMPAGE libraries were constructed from 5ug total RNA following published protocol (Batut and Gingeras 2013) using RiboMinus Plant depletion. shRNAseq libraries were constructed from 5ug of total RNA using TruSeq Small RNA protocol after RiboMinus Plant depletion and size selection of 100 to 205nt with BluePippin (Sage Science).
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| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
Adapter trimming with cutadapt
For RNAseq and RAMPAGE: mapping to W22 v2 genome with STAR (parameters: --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 999 --outFilterMismatchNoverReadLmax 0.04 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --outFilterMultimapNmax 20 --quantMode GeneCounts).
For RAMPAGE only, TSS were identified by calling peak with macs2 using the corresponding RNAseq control.
For shRNA, trimmed fastq files were depleted of snoRNAs, rRNAs and tRNAs by mapping to these structural RNAs from zea mays with bowtie2, and the leftover reads were mapped to W22 v2 with ShortStack (--mmap u --dicermin 20 --dicermax 24 --bowtie_m all --mismatches 1 --foldsize 1000 --pad 250)
Assembly: W22 v2
Supplementary files format and content: For RNAseq: count table on all W22 v2 genes for each sample (combined ReadsPerGene.out.tab files from STAR)
Supplementary files format and content: For RAMPAGeE: list of identified TSS (macs2 narrowPeak format)
Supplementary files format and content: For shRNA: table of identified clusters (ShortStack Results file)
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| Submission date |
May 21, 2024 |
| Last update date |
May 22, 2024 |
| Contact name |
Jonathan Cahn |
| E-mail(s) |
[email protected]
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| Organization name |
CSHL
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| Street address |
1 Bungtown Rd
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| City |
Cold Spring Harbor |
| State/province |
NY |
| ZIP/Postal code |
11724 |
| Country |
USA |
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| Platform ID |
GPL20156 |
| Series (2) |
| GSE254489 |
MaizeCODE reveals bi-directionally expressed enhancers that harbor molecular signatures of maize domestication [W22 RNA] |
| GSE254496 |
MaizeCODE reveals bi-directionally expressed enhancers that harbor molecular signatures of maize domestication |
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| Relations |
| BioSample |
SAMN09291126 |
| SRA |
SRX4149424 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8285432_W22_roots_shRNA_Rep2_results.txt.gz |
3.7 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
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