|
Status |
Public on Jan 07, 2022 |
Title |
TbPOLIE RNAi uninduced (-D) rep 1 |
Sample type |
SRA |
|
|
Source name |
culcured cells
|
Organism |
Trypanosoma brucei brucei |
Characteristics |
strain: lister 427 genotype/variation: TbPOLIE+Myc/+ RNAi genotype/variation: TbPOLIE-myc (endogenous tagging) treatment: -doxycycline
|
Treatment protocol |
100 ng/ml doxycycline (to induce TbPOLIE RNAi) for 30 hrs
|
Growth protocol |
Bloodstream T. brucei cells were cultured in HMI9 medium with 10% FBS. All cells were maintained with medium containing 2 µg/ml G418, 4 µg/ml Hygromycin, and 2 µg/ml of Phleomycin.
|
Extracted molecule |
polyA RNA |
Extraction protocol |
Total RNA was isolated and purified through RNeasy columns (Qiagen). RNA samples were run on a BioAnalyzer 2100 (Agilent Technologies) using the Agilent RNA 6000 nano kit to verify the RNA quality before sent to Novogene, where RNA degradation and contamination were monitored on 1% agarose gels. RNA purity was checked using the NanoPhotometer® spectrophotometer (IMPLEN). RNA integrity and quantitation were assessed using the RNA Nano 6000 Assay Kit on a Bioanalyzer 2100 system (Agilent Technologies). Sequencing libraries were generated using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB, USA) using 1 μg poly(A) RNA according to manufacturer’s protocol and index codes were added to attribute sequences to each sample.
|
|
|
Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina NovaSeq 6000 |
|
|
Data processing |
Base-calling using NovaSeq 6000 for Illumina sequencing Raw reads were first processed with Novogene Perl Scripts (removing reads with low quality, with adapters, and containing poly-N; calculating Q20, Q30 and GC contents). Clean reads were mapped to T. brucei Lister 427 genome TriTrypDB-45_TbruceiLister427_2018_Genome.fasta and its annotation TriTrypDB-45_TbruceiLister427_2018.gff Idownloaded from the TriTryp DB) using HISAT2. HTSeq v0.6.1 was used to count the read numbers mapped to each gene. FPKM of each gene was calculated based on the length of the gene and reads count mapped to this gene Differential expression analysis of two conditions/groups (three biological replicates per condition) was performed using the DESeq R package (1.18.0)
|
|
|
Submission date |
Aug 27, 2021 |
Last update date |
Jan 07, 2022 |
Contact name |
Bibo Li |
E-mail(s) |
b.li37@csuohio.edu
|
Phone |
216 6872444
|
Organization name |
CLEVELAND STATE University
|
Department |
Center for Gene Regulation in Health and Disease
|
Street address |
2121 Euclid Avenue
|
City |
CLEVELAND |
State/province |
OH |
ZIP/Postal code |
44115 |
Country |
USA |
|
|
Platform ID |
GPL27986 |
Series (1) |
GSE182941 |
POLIE suppresses telomerase-mediated telomere G-strand extension and helps ensure proper telomere C-strand synthesis in trypanosomes |
|
Relations |
BioSample |
SAMN21024289 |
SRA |
SRX11952362 |