|
| Status |
Public on Feb 16, 2016 |
| Title |
1µM AGE-BSA |
| Sample type |
SRA |
| |
|
| Source name |
ligament cells_stimulated
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Primary human posterior longitudinal ligament cells
stimulated with: 1µM AGE-BSA
passage: 3
|
| Treatment protocol |
AGE-BSA (1µM or 5µM) or BSA with or without 5µM of BMP2 were added to the medium, and cells were collected for further analysis after 3 days of treatment
|
| Growth protocol |
Normal posterior longitudinal ligament were dissected during the surgery, all tissues were obtained with written informed consent signed by the donors voluntarily for research. Ligaments were further cut into pieces and cultured in DMEM supplemented with 20% fetal bovine serum, and cells of passage 3 were used for the experiments.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from cell or animal tissue by Trizol reagent (Invitrogen) separately. The RNA quality was checked by Bioanalyzer 2200 (Aligent) and kept at -80℃.The RNA with RIN >8.0 is right for cDNA library construction.
The complementary DNA (cDNA) libraries for single-end sequencing were prepared using Ion Total RNA-Seq Kit v2.0 (Life Technologies) according to the manufacturer’s instructions. The cDNA libraries were then processed for the Proton Sequencing process according to the commercially available protocols. Samples were diluted and mixed, the mixture was processed on a OneTouch 2 instrument (Life Technologies) and enriched on a OneTouch 2 ES station (Life Technologies) for preparing the template-positive Ion PI™ Ion Sphere™ Particles (Life Technologies) according to Ion PI™ Template OT2 200 Kit v2.0 (Life Technologies). After enrichment, the mixed template-positive Ion PI™ Ion Sphere™ Particles of samples was loaded on to 1 P1v2 Proton Chip (Life Technologies) and sequenced on Proton Sequencers according to Ion PI Sequencing 200 Kit v2.0 (Life Technologies).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Ion Torrent Proton |
| |
|
| Data processing |
The complementary DNA (cDNA) libraries for single-end sequencing were prepared using Ion Total RNA-Seq Kit v2.0 (Life Technologies) according to the manufacturer’s instructions. The cDNA libraries were then processed for the Proton Sequencing process according to the commercially available protocols. Samples were diluted and mixed, the mixture was processed on a OneTouch 2 instrument (Life Technologies) and enriched on a OneTouch 2 ES station (Life Technologies) for preparing the template-positive Ion PI™ Ion Sphere™ Particles (Life Technologies) according to Ion PI™ Template OT2 200 Kit v2.0 (Life Technologies). After enrichment, the mixed template-positive Ion PI™ Ion Sphere™ Particles of samples was loaded on to 1 P1v2 Proton Chip (Life Technologies) and sequenced on Proton Sequencers according to Ion PI Sequencing 200 Kit v2.0 (Life Technologies).
The clean reads were then aligned to human genome (version: hg19) using the MapSplice program (v2.1.6). In alignment, preliminary experiments were performed to optimize the alignment parameters (-s 22 -p 15 --ins 6 --del 6 --non-canonical) to provide the largest information on the AS events.
We applied HTseq to calculate the mRNA counts based on the mapping result
Genome_build: hg19
Supplementary_files_format_and_content: fastq file and txt file
|
| |
|
| Submission date |
Feb 16, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Chen Xu |
| E-mail(s) |
chenxu8836@hotmail.com
|
| Phone |
+86-13774294166
|
| Organization name |
The Second Military Medical University
|
| Street address |
No, 800, Rd. Xiangyin
|
| City |
Shanghai |
| ZIP/Postal code |
200433 |
| Country |
China |
| |
|
| Platform ID |
GPL17303 |
| Series (1) |
| GSE65952 |
Transcriptome analysis of Advanced glycation end products treated ligament cells |
|
| Relations |
| BioSample |
SAMN03349682 |
| SRA |
SRX878524 |
