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| Status |
Public on Oct 01, 2010 |
| Title |
Pico profiling from 10 cells |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by array
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| Summary |
Current expression profiling methods use RNA from hundreds of thousands or thousands cells. Many fields of biology can not use microarrays due to the nature of the biological systems used that are formed by hundreds or dozens of cells. Here we present a method that can handle RNA amount limitation and gives gene expression profiles from as little as 10 cells. We first validate the method hybridizing amplified RNA from MAQC samples A and B. To do that, 25 ng or 100 pg were used and expression profiles obtained as good as when compared to Affymetrix's chemistry for amplification and labeling. The same experiment was done but using sorted cells from two comercial cell lines (SW620 and SW480) obtaining the same differential expression profiling from 2000 cells or 10 cells. The central step of the method is Whole Transcriptome Amplification (WTA) from Sigma that allows the amplification of very small amounts of RNA as starting material.
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| Overall design |
MAQC samples A and B where amplified using Whole Transcriptome Amplification (WTA) kit from Sigma starting from 25 ng (standard conditions) or 100 pg in triplicates. Amplified cDNA was the fragmented and labeled using the 10kv2 SNP chemistry from Affymetrix and hybridized onto Affymetrix GeneST arrays. The same method was used for RNA isolated from 2000 and 10 cells from cell lines SW620 and SW480. RNA was isolated by magnetic bead purification after treating sorted cells with high DTT concentration and 65ºC to inactivate RNAses.
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| Contributor(s) |
Auer H, Gonzalez E, Rodriguez-Mulero S |
| Citation(s) |
21203435 |
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| Submission date |
Mar 02, 2010 |
| Last update date |
Jul 26, 2018 |
| Contact name |
Victoria Core |
| E-mail(s) |
[email protected]
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| Organization name |
Institute for Research in Biomedicine
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| Department |
Functional Genomics Core
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| Street address |
Baldiri Reixac, 10
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| City |
Barcelona |
| ZIP/Postal code |
08028 |
| Country |
Spain |
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| Platforms (1) |
| GPL6244 |
[HuGene-1_0-st] Affymetrix Human Gene 1.0 ST Array [transcript (gene) version] |
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| Samples (24)
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| GSM517282 |
Human Universal Reference RNA, 25 ng, replicate 1 |
| GSM517283 |
Human Universal Reference RNA, 25 ng, replicate 2 |
| GSM517284 |
Brain RNA, 25 ng, replicate 1 |
| GSM517285 |
Brain RNA, 25 ng, replicate 2 |
| GSM517286 |
Human Universal Reference RNA, 25 ng, replicate 3 |
| GSM517287 |
Brain RNA, 25 ng, replicate 3 |
| GSM517288 |
Human Universal Reference RNA, 100 pg, replicate 1 |
| GSM517289 |
Human Universal Reference RNA, 100 pg, replicate 2 |
| GSM517290 |
Human Universal Reference RNA, 100 pg, replicate 3 |
| GSM517291 |
Brain RNA, 100 pg, replicate 1 |
| GSM517292 |
Brain RNA, 100 pg, replicate 2 |
| GSM517293 |
Brain RNA, 100 pg, replicate 3 |
| GSM517294 |
cell line SW620, 2000 cells, replicate A |
| GSM517295 |
cell line SW620, 2000 cells, replicate C |
| GSM517296 |
cell line SW620, 2000 cells, replicate D |
| GSM517297 |
cell line SW480, 2000 cells, replicate A |
| GSM517298 |
cell line SW480, 2000 cells, replicate C |
| GSM517299 |
cell line SW480, 2000 cells, replicate D |
| GSM517300 |
cell line SW620, 10 cells, replicate A |
| GSM517301 |
cell line SW620, 10 cells, replicate B |
| GSM517302 |
cell line SW620, 10 cells, replicate C |
| GSM517303 |
cell line SW480, 10 cells, replicate A |
| GSM517304 |
cell line SW480, 10 cells, replicate B |
| GSM517305 |
cell line SW480, 10 cells, replicate C |
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| Relations |
| BioProject |
PRJNA124831 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE20595_RAW.tar |
111.5 Mb |
(http)(custom) |
TAR (of CEL) |
| Processed data included within Sample table |
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