|
| Status |
Public on Nov 10, 2013 |
| Title |
Hybrid_RNASeq_02 |
| Sample type |
SRA |
| |
|
| Source name |
Cell
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain: BY4741xRM11-1alpha
cell type: a/alpha diploid
|
| Growth protocol |
Overnight subcultures were used to prepare four technical replicates each of BYxRM hybrid (WL201) cultures and BY+RM co-cultures with starting OD600 at 0.1 and harvested overnight subcultures were used to prepare four technical replicates each of hybrid (WL201) cultures and BY+RM co-cultures with starting OD600 at 0.1 and harvested when cell density reached OD600=1.0 at 30'C, 250rpm
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Each of the replicated subcultures of BYxRM hybrid and BY+RM co-culture harvested at OD600=1.0 was used to extract total RNA by the hot acid phenol method (Kohrer and Domdey 1991)
mRNA-seq library preparation used the Illumina TruSeq mRNA-seq Sample Prep kit with some modifications. Briefly, 5 μg of total RNA from each sample was used to purify for polyA-RNA, and the mRNA was fragmented by heat at 94℃ for 8 min. Double-stranded cDNA was synthesized by random priming, end repaired, and ligated to the Y-shaped TruSeq adaptors. Samples were cleaned by AMPure beads (Agencourt) and split into two halves, which were independently assembled into two PCR reactions, one using the PCR reagents provided in the Illumina kit, and the other using the KAPA PCR reagent (KAPA HiFi HotStart ReadyMix). All reactions went through 12 cycles of PCR amplifications, and were cleaned by AMPure beads to remove primer dimers. The purified products were quantified by Qubit (Invitrogen) and BioAnalyzer 2100 High Sensitivity DNA Assay (Agilent).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer IIx |
| |
|
| Description |
a/alpha diploid
|
| Data processing |
Illumina CASAVA 1.8.2 software used for basecalling
RNA-seq reads were analyzed by ASAP (“Allele-Specific Alignment Pipeline”). To determine whether a given sequence matches one of the two reference genomes specifically, it performs alignments against both sequences in parallel using the Bowtie program (Langmead et al. 2009). In our analysis, the seed length was set to 40 and the maximum number of mismatches permitted in the seed was set to 2.
Genome_build: SGD (version R58-1-1) & RM11-la Database (BROAD Inst.)
Supplementary_files_format_and_content: each tab-delimited text file includes counts of reads mapped on SNPs of each of the orthologous gene pairs
|
| |
|
| Submission date |
May 10, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Li-Ching Hsieh |
| Organization name |
Academia Sinica
|
| Department |
Biodiversity Research Center
|
| Lab |
Li Lab
|
| Street address |
128 Academia Road, Section 2
|
| City |
Taipei |
| ZIP/Postal code |
115 |
| Country |
Taiwan |
| |
|
| Platform ID |
GPL13272 |
| Series (1) |
| GSE46838 |
Inheritance of gene expression level and selective constraints on trans- and cis-regulatory changes in yeast |
|
| Relations |
| BioSample |
SAMN02142059 |
| SRA |
SRX277162 |
