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| Status |
Public on Feb 18, 2015 |
| Title |
AF44 |
| Sample type |
SRA |
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| Source name |
AF44
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| Organism |
Methanosarcina acetivorans C2A |
| Characteristics |
strain: WWM833
genotype: Dhpt DmsrH
growth medium: High Salt medium with 5 mM TMA supplemented with 20 mM MMPA
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| Growth protocol |
cells were grown to mid-exponential phase
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| Extracted molecule |
total RNA |
| Extraction protocol |
The total RNA was isolated from early exponential phase cultures using TRIzol (Invitrogen, Carslbad, CA) and Zymo Direct-zol RNA MiniPrep kits (Zymo Research, Irvine, CA). The 16S- and 23S-rRNA were subtracted using sample-specific biotin-labeled 16S and 23S RNA probes
Fifty nanograms of rRNA-depleted messenger RNA were converted to indexed RNAseq libraries with the ScriptSeq™ v2 RNA-Seq Library Preparation Kit (Epicentre Biotechnologies, Madison, WI). The libraries were pooled in equimolar concentration and quantitated by qPCR with the Library Quantification kit Illumina compatible (Kapa Biosystems, Woburn, MA). The pooled libraries were sequenced for 101 cycles plus 7 cycles for the index read on a HiSeq2000 using TruSeq SBS version 3 reagents. The fastq files were generated and demultiplexed with Casava 1.8.2 (Illumina, San Diego, CA).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
AF44_GGCTAC_L007_R1_001
TableS2_GEO.xlsx
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| Data processing |
CLC Genomics Workbench (version 7.0)( Aarhus, Denmark) is used to perform all subsequent bioinformatic analysis.
Briefly, fastq files were imported as high-throughput data, and trimmed for quality (Quality: 0.001, Ambiguous: 2, Discard: min 30, max 100) and to remove adapter sequences.
After trimming, remaining reads that mapped to stable RNAs (16s, 23s, 5s rRNA and all tRNAs) were bioinformatically removed (similarity: 0.9, length: 0.85, mismtach: 2, insertion: 3, deletion: 3; global alignment and stand-alone read).
Remaining reads were then mapped to the genome for RNA-seq analyses (mapping parameters: 0.9, length: 0.85, mismatch: 2; insertion: 3; deletion: 3; maximum number of hits for a read: 10).
Expression analysis was set up in CLC before the data were normalized and statistical analysis were done using empirical analysis of DGE (digital gene expression), with total counter filter cutoff set as 5.0. From here, the xlsx files were generated.
genome build: ASM734v1
Supplementary_files_format_and_content: Excel files (xlsx) include normalized mean values for each sample
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| Submission date |
Dec 19, 2014 |
| Last update date |
Oct 28, 2022 |
| Contact name |
He Fu |
| E-mail(s) |
[email protected]
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| Organization name |
University of Georgia
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| Department |
Microbiology
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| Street address |
120 Cedar St
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| City |
Athens |
| State/province |
GA |
| ZIP/Postal code |
30602 |
| Country |
USA |
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| Platform ID |
GPL19569 |
| Series (1) |
| GSE64349 |
High-throughput RNA sequencing of methanosarcina grown on methylated sulfur compounds |
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| Relations |
| BioSample |
SAMN03269163 |
| SRA |
SRX818476 |
| Supplementary data files not provided |
SRA Run Selector |
| Processed data are available on Series record |
| Raw data are available in SRA |
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