|
| Status |
Public on May 08, 2022 |
| Title |
Os18SrRNA Root_DMS(-) |
| Sample type |
SRA |
| |
|
| Source name |
rice roots
|
| Organism |
Oryza sativa Japonica Group |
| Characteristics |
tissue: roots
age: Three-week-old
|
| Treatment protocol |
Three-week-old rice seedlings were collected and immersed in 20ml of 1X DMS reaction buffer (40mM HEPES pH7.5, 100mM KCl and 0.5mM MgCl2). 200, 400, 500µl of DMS was added to the reaction buffer to give a final concentration of 1%, 2% and 2.5% , respectively.The same volume of DEPC-treated water was added into the reaction buffer for the mock treatment. DMS treatment was performed for 15 min at 30°C with shaking at 250rpm or under vacuum (approximately 12 psi) at room temperature without shaking. To quench the reaction, 6 ml β-Mercaptoethanol was added to a final concentration of 23%, and the mixture was incubated for 5 min under vacuum.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
DMS-treated or DMS-Untreated samples were pelleted and resuspended in 1 mL Trizol, RNA was extracted by phenol-chloroform extraction and isopropanol precipitation.rRNA was subtracted with Ribo-zero rRNA removal kit for plant (illumina).
For target-specific DMS-MaPseq, 6 µg of DMS-treated sample was mixed with gene-specific RT primers. The reverse transcription was proceeded with TGIRT-III (Ingex). Then specific gene cDNA were amplified using KOD hot start DNA polymerase with gene-specific primers. For genome-wide DMS, DMS-MaPseq library was constructed using Illumina TruSeq® Stranded Total RNA Sample Prep Plant kit (Illumina) and TGIRT enzyme (InGex).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
18SrRNA
|
| Data processing |
Adapters were trimmed by TrimGalore, clean reads over 70bp were retained.
Reads were aligned using Tophat allowing 10% mismatches. Next, uniquely mapped reads were extracted using the Linux command grep with NH:I:1 tag.
Mismatch count and coverage in each nuclotide were called, and mismatches located within 3nt of an insertions or deletion were masked for future analysis.
Assembly: MSU Rice Genome Annotation Project Release 7
Supplementary files format and content: text files include mismatch count and coverage in each nucleotide for each Sample.
|
| |
|
| Submission date |
May 05, 2022 |
| Last update date |
May 08, 2022 |
| Contact name |
Xiaorong Fan |
| E-mail(s) |
[email protected]
|
| Organization name |
Nanjing Agricultural University
|
| Department |
1State Key Laboratory of Crop Genetics and Germplasm Enhancement
|
| Street address |
Nanjing city Wei Tong Road 6
|
| City |
Nanjing |
| ZIP/Postal code |
210095 |
| Country |
China |
| |
|
| Platform ID |
GPL27860 |
| Series (1) |
| GSE197245 |
Probing in vivo RNA structure with optimized DMS-MaPseq in rice |
|
| Relations |
| BioSample |
SAMN28102233 |
| SRA |
SRX15165819 |
