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| Status |
Public on Jan 24, 2024 |
| Title |
m6Apool2_Input_ESC_CAST-HOUS_rep1 |
| Sample type |
SRA |
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| Source name |
mice cell lines ESC hybrid
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| Organism |
Mus musculus |
| Characteristics |
cell line: ESC hybrid
treatment: m6A-seq2
fraction: input
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| Growth protocol |
All yeast strains used in this work were grown in YPD (2% dextrose) at 30C. To induce synchronous sporulation, cells were grown for 24 h in 1% yeast extract, 2% peptone, 4% dextrose at 30C, diluted in BYTA (1% yeast extract, 2% tryptone, 1% potassium acetate, 50 mM potassium phthalate) to OD600 = 0.2 and grown for another 16 h at 30C, 250 rpm. Cells were then washed two times with water and re-suspended in Sporulation medium (0.3% potassium acetate) at OD600 = 2.0 and incubated at 30C at 190 rpm. Human monochromosome hybrid cells JCRB2201, JCRB2202 and JCRB2203 were purchased from the Japanese Collection of Research Bioresources (JCRB), and cultured in DMEM (GIBCO) supplemented with 10% fetal bovine serum (FBS) and 0.8mg/ml G418(Gibco,79N9555) 37. mESCs, BJ, and NIH 3T3 cells were cultured in DMEM (GIBCO) supplemented with 10% FBS and 1% Penicillin and Streptomycin.
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
Yeast total RNA samples were prepared by adjusting previously published protocols. After fast freezing the yeast cells in liquid nitrogen, yeast cells were resuspended in equal amounts of phenol:Chloroform:Isoamyl alcohol (Sigma Aldrich), buffer AE (50mM sodium acetate, 10mM EDTA 1% SDS), and glass beads in Eppendorf tube. The tube was vortexed for 15 minutes in a bullet blender, heated to 65 °C for 30 minutes, followed by another round of 5 minutes vortex and 30 minutes heating to 65 °C. Samples were placed on ice for 5 minutes and centrifuged for 10 minutes (12000g, 4 °C). The supernatant was isolated, re-extracted with phenol:Chloroform:Isoamyl alcohol, and precipitated with sodium acetate and isopropanol. For all types of mammalian cell lines, total RNA was extracted with BIO TRI RNA reagent (Bio-lab). Enrichment for mRNA was done by two rounds of poly-A selection using Oligo(dT) beads (Dynabeads mRNA DIRECT) in both total RNAs from yeast and mammals.
m6A-seq2: Fragmentation, 3' adapter ligation, pooled(m6A-IP, cDNA synthesis, second adapter ligation and enrichment)
m6A-seq2
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| Library strategy |
RIP-Seq |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
alignment using STAR (V. 2.5.3a)
Read coverages were obtained using the R package txtools (https://github.com/AngelCampos/txtools)
Assembly: SK1-YPS138, hg19-mm9, CAST-HOUS
Supplementary files format and content: rds
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| Submission date |
May 12, 2023 |
| Last update date |
Jan 24, 2024 |
| Contact name |
Ran Shachar |
| E-mail(s) |
ran.shachar@weizmann.ac.il
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| Organization name |
Weizmann Institute of Science
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| Department |
Department of Molecular Genetics
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| Street address |
234 Herzl St.
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| City |
Rehovot |
| ZIP/Postal code |
7610001 |
| Country |
Israel |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE232450 |
Dissecting the sequence and structural determinants guiding m6A deposition and evolution via inter- and intra-species hybrids |
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| Relations |
| BioSample |
SAMN35065553 |
| SRA |
SRX20314118 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7336553_m6Apool2_miceHYBRID_IN_rep1_Aligned.out.sorted.txDT.rds.gz |
67.4 Mb |
(ftp)(http) |
RDS |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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