|
| Status |
Public on Jun 13, 2013 |
| Title |
N48_CD10+_SAGEseq |
| Sample type |
SRA |
| |
|
| Source name |
human CD10+ breast myoepithelial cells
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: breast
cell type: bead purified CD10+ myoepithelial cells
parity: nulliparous
|
| Growth protocol |
Primary cells purified from normal human breast tissue
|
| Extracted molecule |
total RNA |
| Extraction protocol |
For SAGE-seq: mRNA isolated from atleast 50-100K cells were captured on oligo(dT) beads followed by cDNA synthesis. cDNA was then cleaved with anchoring enzyme NlAIII following ligation of adapter 1. cDNA was subsequently released by tagging enzyme MmeI and ligated to adapter 2. The resulting 85 bp DNA fragments were amplified by Primer 1 and Primer 2 and purified in 12% PAGE. Purified libraries were sequenced on the Illumina Solexa 1G platform. All the adapters and primers were from Illumina.
For MSDK-seq: 1-2ug of DNA was digested with methylation sensitive restriction enzyme BssHII (NEB) and ligated to biotinylated linker. DNA was then fragmented with another restriction enzyme NlaIII (NEB) and the resulting fragmented DNA with biotinylated linker was captured on streptavidin beads. Fragmented DNA was then ligated to Adapter 1 followed by release of DNA from beads by tagging enzyme MmeI (NEB) with subsequent ligation to Adapter 2. The resulting 85 bp DNA fragments were amplified by Primer 1 and Primer 2 and purified in 12% PAGE. Purified libraries were sequenced on the Illumina Solexa 1G platform. All the adapters and primers were supplied by Illumina.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer |
| |
|
| Description |
SAGE_library
|
| Data processing |
21 base pair (bp) of each read is used in this study. The first 4 bp is always CATG, as this is the recognition site of NlaIII. Our analysis was based around the alignment of the rest of the 17bp tags to human genome composed of segments between nearest NlaIII sites flanking BssHII recognition sites where as 17 bp SAGE tags were aligned to transcripts.
|
| |
|
| Submission date |
Sep 09, 2011 |
| Last update date |
May 15, 2019 |
| Contact name |
Kornelia Polyak |
| E-mail(s) |
[email protected]
|
| Phone |
617-632-2106
|
| Organization name |
Dana-Farber Cancer Institute
|
| Department |
Medical Oncology
|
| Lab |
Polyak
|
| Street address |
450 Brookline Ave
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02215 |
| Country |
USA |
| |
|
| Platform ID |
GPL9052 |
| Series (1) |
| GSE32017 |
Molecular profiling of human mammary gland links breast cancer risk to a p27+ cell population with progenitor characteristics |
|
| Relations |
| Reanalyzed by |
GSE113795 |
| SRA |
SRX097131 |
| BioSample |
SAMN00717463 |
