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| Status |
Public on Nov 06, 2018 |
| Title |
BH500(pSW4)_OD=3_Run1 |
| Sample type |
SRA |
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| Source name |
BH500(pSW4)_OD=3
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| Organism |
Bacillus anthracis str. Ames |
| Characteristics |
strain: BH500
growth phase: lag phase
protocol: control
expression: Non-Producer
od at harvest: 3
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| Extracted molecule |
total RNA |
| Extraction protocol |
Culture samples were immediately treated with RNA protect and preserved in Trizol at -80 degree celcius and total RNA was extracted using hot phenol method.
The NEBNext® Ultra™ RNA Library Prep Kit for Illumina was used to prepare the complete RNA-seq library preparation. First strand cDNA synthesis was performed using NEBNext first strand synthesis reaction buffer, random primers, Protoscript II reverse. To the same reaction mix, second strand synthesis enzyme mix was added and second strand synthesis was. The synthesized cDNA strands were purified using a 1:1 ratio of Agencourt AMPure XP beads (Beckman Coulter Genomics, Danvers, MA). Later steps involved end repair of extracted cDNA, adapter ligation using blunt/TA ligase and cleavage by USER enzyme, purification with AMPure XP beads and PCR enrichment of cDNA library using different sets of Multilex oligos for Illumina. The enriched library was again purified with Agencourt AMPure XP beads, and its quality was checked on a Bioanalyzer using Agilent DNA high sensitivity chips according to the manufacturer’s instructions.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
pSW41OD3
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| Data processing |
Bcl2fastq2 software was used for base-calling.
The sequenced reads were trimmed to remove the low quality bases with the following parameters Trim based on : Quality score; Min read length : 25; Max N : FALSE ; End min quality level (Phred) : 20 ; Trim from end : 3-prime (right end).
Bowtie2 method was used to align the RNA-seq reads with the reference organism Bacillus anthracis str ames AE017334.2.
The quantification of transcriptome abundance was performed using Partek E/M algorithm in PartekFlow software.
Data was normalized using TPM method and differential gene expression were predicted across comparisions using ANOVA.
Genome_build: ASM844v1
Supplementary_files_format_and_content: tab-delimited text file contains normalized values obtained after TPM normalization, addition +1 and log2 transformation.
tab-delimited text file contains results from ANOVA and includes fold change calculations among pYS5 LagPhase vs pSW4 LagPhase, pYS5 LogPhase vs pSW4 LogPhase, pYS5 LateLogPhase vs pSW4 LateLogPhase, along with p-value, FDR stepup etc.
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| Submission date |
Jan 09, 2018 |
| Last update date |
Nov 06, 2018 |
| Contact name |
Ashish K Sharma |
| E-mail(s) |
ashish.sharma@nih.gov, ashish-sharma@idexx.com, ashish.kumar07@gmail.com
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| Phone |
3014022981
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| Organization name |
National Institute of Health
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| Department |
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
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| Lab |
Biotechnology Core
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| Street address |
14 Service Rd West Building 14A Room 176 MS-5522
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| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
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| Platform ID |
GPL24488 |
| Series (1) |
| GSE108973 |
Global transcriptomic response of modified Bacillus anthracis over expressing protective antigen |
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| Relations |
| BioSample |
SAMN08333558 |
| SRA |
SRX3545445 |
| 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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