|
| Status |
Public on Jun 17, 2020 |
| Title |
Ty-I |
| Sample type |
SRA |
| |
|
| Source name |
Parasite cell culture
|
| Organism |
Trypanosoma cruzi |
| Characteristics |
strain: Y strain
cell type: T.cruzi Trypomastigotes
treatment: control
chip antibody: none
|
| Treatment protocol |
The commercially available extracellular matrix (Geltrex ™ LDEV-Free Reduced Growth Factor Basement Membrane Matrix, Gibco, 150 μl ECM per sample) was used to incubate part of the T cruzi trypomastigotes (5 x 10^8 cells). After incubation, the parasites were collected and used for chromatin extraction (Sample named MTy). As a control, 5 x 10^8 trypomastigotes from the same culture were subjected to the same conditions mentioned above in the absence of ECM (sample named Ty). The samples were frozen at -80°C for further use.
|
| Growth protocol |
Cell culture of T. cruzi trypomastigotes (Y strain) were maintained by infection of LLC-MK2 cells in Minimum Essential Medium (MEM – GIBCO®) containing 2% of Fetal Bovine Serum (FBS), under 5% CO2 and 37oC. Five days after infection of the host cell, culture supernatants containing the trypomastigotes were centrifuged and purified.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
After cell lysis in 3 steps, the samples were clarified from sonicated nuclei and Protein-DNA complexes were isolated with antibody.
The libraries of the immunoprecipitated DNA fragments from each sample and the corresponding INPUTs were constructed using the TruSeq ChIP Sample Prep Kit (Illumina, IP-202-1012), following the manufacturer's recommendations.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
First step was the avaluation of quality of the reads sequenced using FASTQC tool.
ChIP-seq reads were aligned to the T cruzi CL-Brener Esmeraldo-like genome (TriTrypDB, release 9) using BWA v 0.7.15-1140, "bwa aln" default parameters. Bwa SAMPE was used to group the files containing the pairs of reads and Samtools was used to obtain the final file properly filled in BAM format.
A Phred mapping quality filter (MAPQ) of 30 was implemented to select the uniquely mapped reads in the T. cruzi genome.
Peaks were called using MACS v2.2 with two distinct sets of parameters for identification of narrow and broad regions. The parameter "--keep-dup auto", using cutoff value of 1e-5 was used to remove part of duplicated reads. The GEM-Mappability Calculator was used to calculate the effective genome size used in MACS2.
The DiffBind v2.2.12 was used to determine the differential profiles of enrichment in each treatment, TY and MTY considering "False Discovery Rate" (FDR) lower than 5%. The statistical analysis is done using DESeq 2 model.
Genome_build: T. cruzi CL-Brener Esmeraldo-like (TritrypDB, release 9)
Supplementary_files_format_and_content: Bedgraph files was generate from MACS2 (Treat and lambda control).
|
| |
|
| Submission date |
Nov 12, 2019 |
| Last update date |
Jun 17, 2020 |
| Contact name |
Rubens Daniel Miserani Magalhães |
| E-mail(s) |
criphg@gmail.com
|
| Organization name |
Ribeirão Preto Medical School
|
| Department |
Department of Cellular and Molecular Biology and Pathogenic Bioagents
|
| Lab |
Cellular and Molecular Parasitology
|
| Street address |
Av. Bandeirantes 3900
|
| City |
Ribeirão Preto |
| State/province |
SP |
| ZIP/Postal code |
14049-900 |
| Country |
Brazil |
| |
|
| Platform ID |
GPL27740 |
| Series (1) |
| GSE140245 |
Global changes in nitration levels and DNA binding profile of Trypanosoma cruzi histones induced by incubation with host extracellular matrix |
|
| Relations |
| BioSample |
SAMN13264261 |
| SRA |
SRX7135167 |
