|
| Status |
Public on Aug 18, 2016 |
| Title |
CD34_cel-2 |
| Sample type |
RNA |
| |
|
| Source name |
CD34 cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: CD34+ human hematopoietic progenitor cells
sample type: cells
|
| Growth protocol |
Human CD34+ cord-blood progenitors (New York Blood Center) were enriched using MACS cell separation (Miltenyi Biotec) and cultured in serum-free media (Stem Cell Technologies) supplemented with 100U/ml penicillin/streptomycin, 40ng/ml FLT3L, 25ng/ml SCF, and 50ng/ml TPO (Miltenyi Biotec) at 1% oxygen and 5% carbon dioxide for 48h, after which exosomes were isolated by differential centrifugation. HL-60 and MOLM-14 cell lines were cultured at 1% oxygen and 5% carbon dioxide in RPMI + 10% FBS and 1X Pen-Strep for 48 hours, after which extracellular vesicles were isolated by differential centrifugation.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the Qiagen miRNeasy kit according to the manufacturer's instructions.
|
| Label |
biotin
|
| Label protocol |
Biotinylated RNA was prepared using Affymetrix HSR miRNA Flash Tag Biotin protocol with 130 ng total RNA input
|
| |
|
| Hybridization protocol |
Hybridizations were performed as per Affymetrix FlashTag Biotin HSR RNA Labeling Kit User Guide with incubation at 48°C/ 60 rpm for 18 hours. Arrays were washed and stained on Fluidics Station 450 using script FS450_0002
|
| Scan protocol |
Standard Affymetrix scan protocol using GeneChip Scanner 3000 7G
|
| Description |
CD34C_2
|
| Data processing |
Data were analyzed with Partek Genomics Suite(v.6.6) using the RMA normalization method. Four samples from GEO accession GSE55025 (GSM1328048, GSM1328049, GSM1328044, GSM1328045) were included along with twelve new samples for the normalization.
|
| |
|
| Submission date |
Aug 17, 2016 |
| Last update date |
Aug 18, 2016 |
| Contact name |
Noah Hornick |
| E-mail(s) |
hornickn@ohsu.edu
|
| Phone |
503-494-0829
|
| Organization name |
OHSU
|
| Department |
Pediatric Hematology Oncology
|
| Lab |
Peter Kurre
|
| Street address |
3181 S.W. Sam Jackson Park Rd.
|
| City |
Portland |
| State/province |
OR |
| ZIP/Postal code |
97239-3098 |
| Country |
USA |
| |
|
| Platform ID |
GPL16384 |
| Series (1) |
| GSE85769 |
Expression and exosome incorporation of microRNA by leukemia versus nonmalignant hematopoietic progenitors |
|
