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| Status |
Public on Jun 14, 2018 |
| Title |
brpL mutant Rep4 2 |
| Sample type |
SRA |
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| Source name |
brpL_mutant
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| Organism |
Streptococcus sanguinis |
| Characteristics |
strain: parent strain: SK36
growth phase: late-log
genotype: brpL mutant
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| Growth protocol |
Cells were incubated at 37°C under anaerobic conditions in brain-heart infusion media and harvested at late-log phase. The brpL mutant was supplemented with kanamycin (500 µg/ml) for selection of the antibiotic resistance cassette.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were lysed and RNA was extracted by FastPrep lysing matrix beads. Clean-up and DNase I treatment was conducted by QIAGEN RNeasy mini kit, according to the manufacterer's protocol. Illlumina RiboZero Magnetic Kit for bacteria was used to deplete ribosomal RNA from 2 µg of total RNA.
NEBNext Ultra Directional RNA Library Prep Kit for Illumina (New England BioLabs) was used for the RNA-seq library preparation according to the manufacturer’s protocol. Briefly, ribosomal-depleted RNA was fragmented followed by first-strand cDNA synthesis from random primers using ProtoScript II Reverse Transcriptase (New England BioLabs). Second strand cDNA was synthesized and purified. End repair was performed on the double-stranded cDNA and primed with the addition of 5’-phosphorylated dA-tailed ends using T4 DNA polymerase, Klenow DNA polymerase and T4 polynucleotide kinase (New England BioLabs). This was immediately followed by adaptor ligation (New England BioLabs) and purified. Samples were PCR-amplified for 12 cycles with Phusion HiFi polymerase (New England Biolabs) with paired-end primers and a randomly chosen unique barcode (NEB Next Multiplex Oligs for Illumina - Index Primers 2, 11, 12, 13, 15, 16, 18, 20, 21, 22). Agencourt AMPure XP Beads (Beckman Coulter) were used for all purification steps.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
replicate 4
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| Data processing |
Standard Illumina pipeline. Real Time Analysis (RTA v. 1.18.66.3) software used for image processing, intesity extraction and basecalling based on intensities. CHASTITY software used for quality filter of basecalling.
Reads obtained from RNA-sequencing were aligned against the Streptococcus sanguinis SK36 genome using EDGE-pro.
Differential genes analysis was executed by DESeq2.
Genome_build: Streptococcus sanguinis reference genome
Supplementary_files_format_and_content: tab-delimited text file includes gene name and ID, fold change of expression, p-value
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| Submission date |
Feb 07, 2018 |
| Last update date |
Jun 14, 2018 |
| Contact name |
Ping Xu |
| E-mail(s) |
[email protected]
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| Organization name |
Virginia Commonwealth Univ. School of Dentistry
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| Department |
Philips Institute for Oral Health Research
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| Lab |
Ping Xu
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| Street address |
521 North 11th street
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| City |
Richmond |
| State/province |
VA |
| ZIP/Postal code |
23298 |
| Country |
USA |
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| Platform ID |
GPL22689 |
| Series (1) |
| GSE110307 |
Analysis of differential gene expression in the Streptococcus sanguinis brpL mutant |
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| Relations |
| BioSample |
SAMN08481998 |
| SRA |
SRX3660840 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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