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Metadata Template
Matrix non-normalized Template
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Metadata Example
Matrix non-normalized Example
Matrix normalized Example
Fold Change
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| # Use this template for large-scale, high-throughput RT-PCR submissions. | ||||||||||||||||||||
| # In addition to this 'Metadata Template' worksheet, there are related 'Matrix non-normalized', 'Matrix normalized', and 'Fold Change' worksheets. Click tabs below. | ||||||||||||||||||||
| # Most fields in this template must be completed. | ||||||||||||||||||||
| # For 'Example' worksheets, click tabs below. | ||||||||||||||||||||
| # Field names (in blue on this page) should not be edited. Hover over cells containing field names to view field content guidelines or, | ||||||||||||||||||||
| # CLICK HERE for Field Content Guidelines from GEO. | ||||||||||||||||||||
| SERIES | ||||||||||||||||||||
| # This section describes the overall experiment. | ||||||||||||||||||||
| title | ||||||||||||||||||||
| summary | ||||||||||||||||||||
| summary | ||||||||||||||||||||
| overall design | ||||||||||||||||||||
| contributor | ||||||||||||||||||||
| contributor (SEEK ID) | ||||||||||||||||||||
| SEEK Project | Project | |||||||||||||||||||
| Experiment Class (assay Type) | gene_expression_profiling | |||||||||||||||||||
| Experiment Design type | ExperimentDesignType | |||||||||||||||||||
| Technology type | RT-PCR | |||||||||||||||||||
| quality control type | QualityControlDescriptionType | |||||||||||||||||||
| SAMPLES | ||||||||||||||||||||
| # The Sample names in the first column are arbitrary but they must match the column headers of the Matrix table (see next worksheet). | ||||||||||||||||||||
| Sample name | title | description | source name | organism | characteristics:NCBI_ID | characteristics:Strain | characteristics:Genotype | characteristics:Phenotype | characteristics: Tag | molecule | label | platform | ||||||||
| SAMPLE 1 | organism | MaterialType | LabelCompound | |||||||||||||||||
| SAMPLE 2 | organism | MaterialType | LabelCompound | |||||||||||||||||
| SAMPLE 3 | organism | MaterialType | LabelCompound | |||||||||||||||||
| SAMPLE 4 | organism | MaterialType | LabelCompound | |||||||||||||||||
| PROTOCOLS | ||||||||||||||||||||
| # This section includes protocols and fields which are common to all Samples. | ||||||||||||||||||||
| # Protocols which are applicable to specific Samples or specific channels should be included in additional columns of the SAMPLES section instead. | ||||||||||||||||||||
| growth protocol | ||||||||||||||||||||
| treatment protocol | ||||||||||||||||||||
| extract protocol | ||||||||||||||||||||
| label protocol | ||||||||||||||||||||
| hyb protocol | n/a | |||||||||||||||||||
| scan protocol | n/a | |||||||||||||||||||
| data processing | ||||||||||||||||||||
| value definition | ||||||||||||||||||||
| PLATFORM | ||||||||||||||||||||
| # If your panel of PCR assays is already deposited in GEO, please fill in platform column in SAMPLES section and ignore this PLATFORM section. | ||||||||||||||||||||
| # If your panel of PCR assays is not in GEO, please complete the fields below and include platform annotation columns in Matrix normalized Template (see example). | ||||||||||||||||||||
| title | ||||||||||||||||||||
| technology | RT-PCR | |||||||||||||||||||
| distribution | virtual | |||||||||||||||||||
| organism | ||||||||||||||||||||
| manufacturer | ||||||||||||||||||||
| manufacture protocol | see manufacturer's website | |||||||||||||||||||
| catalog number | ||||||||||||||||||||
| description | PCR assay | |||||||||||||||||||
| description | ||||||||||||||||||||
| # Platform Column Definitions: describe the contents of each platform column in the Matrix normalized Template | ||||||||||||||||||||
| header 1 | header 1 definition | |||||||||||||||||||
| header 2 | header 2 definition | |||||||||||||||||||
| header 3 | header 3 definition | |||||||||||||||||||
| etc… | ||||||||||||||||||||
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| # Matrix non-normalized table for RT-PCR submission based on a Platform not yet represented in GEO. | |||||||||||||||||
| # The ID_REF column must contain unique identifiers. | |||||||||||||||||
| # SAMPLE columns should report non-normalized data (e.g., raw Ct_Target). | |||||||||||||||||
| # SAMPLE column header names must match Sample name column in SAMPLES section of Metadata sheet. | |||||||||||||||||
| ID_REF | SAMPLE 1 | SAMPLE 2 | SAMPLE 3 | SAMPLE 4 | |||||||||||||
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| # Matrix normalized table for RT-PCR submission based on a Platform not yet represented in GEO. | |||||||||||||||
| # The ID_REF column must contain unique identifiers. | |||||||||||||||
| # Columns header 1, header 2, header 3, etc., should include Platform annotations. | |||||||||||||||
| # SAMPLE columns should report normalized data used to draw study's conclusions [target gene signal normalized to housekeeping genes; e.g., 2^-ΔCt, where -ΔCt = -(Ct_Target − Ct_HKG)] | |||||||||||||||
| # SAMPLE values that should be disregarded may be left blank or labeled as "null". | |||||||||||||||
| # SAMPLE column header names must match Sample name column in SAMPLES section of Metadata sheet. | |||||||||||||||
| ID_REF | header 1 | header 2 | header 3 | SAMPLE 1 | SAMPLE 2 | SAMPLE 3 | SAMPLE 4 | ||||||||
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| # Fold change data = test/control | |||||||||
| # e.g., SAMPLEtest target gene signal normalized to housekeeping gene (2^-ΔCttest)/SAMPLEcontrol target gene signal normalized to HKG (2^-ΔCtcontrol) | |||||||||
| # 2^-ΔΔCt, where -ΔΔCt = -[ΔCttest -ΔCtcontrol] | |||||||||
| ID_REF | Fold_change 1 | Fold_change 2 | Fold_change 3 | ||||||
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| SERIES | |||||||||||||||||
| title | Real-time quantitative PCR analysis of human dendritic cells | ||||||||||||||||
| summary | Human CD14 positive monocytes were purified from healthy volunteers' blood and were differentiated to immature dendritic cells in vitro by culturing for five days in the presence of interleukin-4 (IL-4 100 ng/ml) and GM-CSF (75 ng/ml). Immature dendritic cells were activated three different ways for 24 hours: 1. Double-stranded DNA poly(dA:dT) (2.5 μg/ml) complexed with LyoVec transfection reagent. 2. LPS (500 ng/ml) 3. Inflammatory cocktail containing 10 ng/ml TNF, 5 ng/ml IL-1β, 20 ng/ml IL-6, 75 ng/ml GM-CSF and 1 μg/ml PGE2. We used SA Biosciences Antigen Presenting PCR assay panel to quantitate gene expression of immunologically relevant genes from the immature and the differently activated cells. | ||||||||||||||||
| overall design | qPCR gene expression profiling. Monocytes from three donors were used and treated separately as indicated in the summary. Equal amount total RNA from each donor was pooled prior to gene expression analysis. | ||||||||||||||||
| contributor | Jane,Doe | ||||||||||||||||
| contributor | John,A,Smith | ||||||||||||||||
| SAMPLES | |||||||||||||||||
| # The corresponding example matrix table is included in the next worksheet. | |||||||||||||||||
| Sample name | title | description | source name | organism | characteristics: cell type | characteristics: activation agent | characteristics: time | molecule | label | platform | |||||||
| SAMPLE 1 | Immature DC | Control | Monocyte-derived immature dendritic cell | Homo sapiens | immature dendritic cell | control | 5 days | total RNA | SYBR Green | ||||||||
| SAMPLE 2 | Double-stranded DNA-activated mature DC | Test | Monocyte-derived immature dendritic cell | Homo sapiens | immature dendritic cell | Double-stranded DNA | 5+1 days | total RNA | SYBR Green | ||||||||
| SAMPLE 3 | LPS-activated mature DC | Test | Monocyte-derived immature dendritic cell | Homo sapiens | immature dendritic cell | LPS | 5+1 days | total RNA | SYBR Green | ||||||||
| SAMPLE 4 | Inflammatory cocktail-activated mature DC | Test | Monocyte-derived immature dendritic cell | Homo sapiens | immature dendritic cell | Inflammatory cocktail | 5+1 days | total RNA | SYBR Green | ||||||||
| PROTOCOLS | |||||||||||||||||
| extract protocol | Total RNA was extracted with RNeasy Mini Kit followed by DNase I treatment. | ||||||||||||||||
| label protocol | PCR assays were performed using the Dendritic Cell - Antigen Presenting Cell and the Toll-like Receptor Pathway PCR assay panel (SA Biosciences, Frederick, MD) following the Manufacturer’s instructions. Reverse transcription was performed from 500 ng total RNA using the RT2 First Strand Kit (SA Biosciences). Quantitative real-time PCR were performed (Light Cycler 480, Roche) with 45 cycles at 94 oC for 15 seconds and 60 oC for 60 seconds. | ||||||||||||||||
| hyb protocol | n/a | ||||||||||||||||
| scan protocol | n/a | ||||||||||||||||
| data processing | The normalization and all the data analysis were performed according to the manufacturers instructions using their web-based software package: http://sabiosciences.com/pcrarraydataanalysis.php | ||||||||||||||||
| data processing | For the normalization it uses the average of five housekeeping genes: B2M, HPRT1 , RPL13A, GAPDH, ACTB. | ||||||||||||||||
| data processing | Target gene signals normalized to housekeeping genes; 2^-deltaCt, where deltaCt = (Ct_Target − Ct_HKG)] | ||||||||||||||||
| data processing | The web-based software package automatically performs all deltadeltaCt based fold-change calculations from the uploaded raw threshold cycle data. | ||||||||||||||||
| value definition | Matrix normalized worksheet reports normalized signal (against housekeeping genes). | ||||||||||||||||
| value definition | Fold Change worksheet reports test/control (i.e., mature DC/immature DC) ratios. | ||||||||||||||||
| PLATFORM | |||||||||||||||||
| # If your panel of PCR assays is already deposited in GEO, please fill in platform column in SAMPLES section and ignore this PLATFORM section. | |||||||||||||||||
| # If your panel of PCR assays is not in GEO, please complete the fields below and include platform annotation columns in Matrix normalized Template (see example). | |||||||||||||||||
| title | SABiosciences Human Dendritic and Antigen Presenting Cell PCR assay panel | ||||||||||||||||
| technology | RT-PCR | ||||||||||||||||
| distribution | virtual | ||||||||||||||||
| organism | Homo sapiens | ||||||||||||||||
| manufacturer | SABiosciences | ||||||||||||||||
| manufacture protocol | see manufacturer's website | ||||||||||||||||
| catalog number | PAHS-406F-12 | ||||||||||||||||
| description | DC-APC PCR assay panel | ||||||||||||||||
| description | SABiosciences RT² Profiler panel of PCR assays. | ||||||||||||||||
| # Platform Column Definitions: describe the contents of each platform column of the matrix table | |||||||||||||||||
| Unigene | UniGene cluster ID | ||||||||||||||||
| GB_ACC | RefSeq accession number of spotted sequence | ||||||||||||||||
| Symbol | Gene symbol | ||||||||||||||||
| Descsription | Gene description | ||||||||||||||||
| Gene Name | Gene name | ||||||||||||||||
| SPOT_ID | controls |
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| # truncated example of Matrix with non-normalized data (raw Ct values for target gene) | |||||||||
| ID_REF | SAMPLE 1 | SAMPLE 2 | SAMPLE 3 | SAMPLE 4 | |||||
| A01 | 35 | 35 | 35 | 35 | |||||
| A02 | 29.35 | 28.19 | 29.63 | 31.63 | |||||
| A03 | 35 | 35 | 35 | 35 | |||||
| A04 | 35 | 35 | 33.08 | 35 | |||||
| A05 | 29.94 | 25.47 | 27.68 | 34.34 | |||||
| A06 | 27.52 | 24.65 | 23.49 | 26.44 | |||||
| A07 | 35 | 31.59 | 29.7 | 35 | |||||
| A08 | 25.09 | 22.52 | 20.6 | 22.83 | |||||
| A09 | 34.37 | 30.67 | 27.49 | 32.26 | |||||
| A10 | 35 | 33.12 | 35 | 35 | |||||
| A11 | 32.2 | 23.76 | 25.71 | 33.92 | |||||
| A12 | 25.48 | 25.26 | 27.05 | 31.17 | |||||
| B01 | 35 | 35 | 35 | 35 | |||||
| B02 | 35 | 35 | 35 | 35 | |||||
| B03 | 29.58 | 28.79 | 30.04 | 35 | |||||
| B04 | 28.04 | 25.92 | 26.52 | 27.87 | |||||
| B05 | 26.75 | 26.84 | 28.15 | 28.77 | |||||
| B06 | 27.3 | 26.84 | 28.26 | 29.07 | |||||
| B07 | 35 | 35 | 35 | 35 | |||||
| B08 | 35 | 35 | 35 | 35 | |||||
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| # truncated example of Matrix with PCR assay annotation columns and normalized target gene signal data | ||||||||||||||||||||||||||||||
| ID_REF | Unigene | GB_ACC | Symbol | Descsription | Gene Name | SPOT_ID | SAMPLE 1 | SAMPLE 2 | SAMPLE 3 | SAMPLE 4 | ||||||||||||||||||||
| A01 | Hs.54460 | NM_002986 | CCL11 | Chemokine (C-C motif) ligand 11 | SCYA11 | 0.003685 | 0.002549 | 0.004187 | 0.004095 | |||||||||||||||||||||
| A02 | Hs.414629 | NM_005408 | CCL13 | Chemokine (C-C motif) ligand 13 | CKb10/MCP-4 | 0.185052 | 0.285983 | 0.173139 | 0.042335 | |||||||||||||||||||||
| A03 | Hs.10458 | NM_004590 | CCL16 | Chemokine (C-C motif) ligand 16 | CKb12/HCC-4 | 0.003685 | 0.002549 | 0.004187 | 0.004095 | |||||||||||||||||||||
| A04 | Hs.50002 | NM_006274 | CCL19 | Chemokine (C-C motif) ligand 19 | CKb11/ELC | 0.003685 | 0.002549 | 0.015843 | 0.004095 | |||||||||||||||||||||
| A05 | Hs.303649 | NM_002982 | CCL2 | Chemokine (C-C motif) ligand 2 | GDCF-2/HC11 | 0.122938 | 1.884263 | 0.668964 | 0.00647 | |||||||||||||||||||||
| A06 | Hs.514107 | NM_002983 | CCL3 | Chemokine (C-C motif) ligand 3 | G0S19-1/LD78ALPHA | 0.657927 | 3.326488 | 12.210074 | 1.545421 | |||||||||||||||||||||
| A07 | Hs.512683 | NM_021006 | CCL3L1 | Chemokine (C-C motif) ligand 3-like 1 | D17S1718/G0S19-2 | 0.003685 | 0.027092 | 0.164938 | 0.004095 | |||||||||||||||||||||
| A08 | Hs.75703 | NM_002984 | CCL4 | Chemokine (C-C motif) ligand 4 | ACT2/G-26 | 3.545527 | 14.560621 | 90.509668 | 18.869705 | |||||||||||||||||||||
| A09 | Hs.514821 | NM_002985 | CCL5 | Chemokine (C-C motif) ligand 5 | D17S136E/RANTES | 0.005703 | 0.051261 | 0.76313 | 0.027356 | |||||||||||||||||||||
| A10 | Hs.251526 | NM_006273 | CCL7 | Chemokine (C-C motif) ligand 7 | FIC/MARC | 0.003685 | 0.009381 | 0.004187 | 0.004095 | |||||||||||||||||||||
| A11 | Hs.271387 | NM_005623 | CCL8 | Chemokine (C-C motif) ligand 8 | HC14/MCP-2 | 0.025666 | 6.164569 | 2.620787 | 0.008657 | |||||||||||||||||||||
| A12 | Hs.301921 | NM_001295 | CCR1 | Chemokine (C-C motif) receptor 1 | CD191/CKR-1 | 2.705696 | 2.179504 | 1.035265 | 0.058234 | |||||||||||||||||||||
| B01 | Hs.511794 | NM_000648 | CCR2 | Chemokine (C-C motif) receptor 2 | CC-CKR-2/CCR2A | 0.003685 | 0.002549 | 0.004187 | 0.004095 | |||||||||||||||||||||
| B02 | Hs.506190 | NM_001837 | CCR3 | Chemokine (C-C motif) receptor 3 | CC-CKR-3/CD193 | 0.003685 | 0.002549 | 0.004187 | 0.004095 | |||||||||||||||||||||
| B03 | Hs.450802 | NM_000579 | CCR5 | Chemokine (C-C motif) receptor 5 | CC-CKR-5/CCCKR5 | 0.157782 | 0.188678 | 0.130308 | 0.004095 | |||||||||||||||||||||
| B04 | Hs.1309 | NM_001763 | CD1A | CD1a molecule | CD1/FCB6 | 0.45882 | 1.379361 | 1.494849 | 0.573554 | |||||||||||||||||||||
| B05 | Hs.1310 | NM_001764 | CD1B | CD1b molecule | CD1/CD1A | 1.121943 | 0.729005 | 0.482968 | 0.30736 | |||||||||||||||||||||
| B06 | Hs.132448 | NM_001765 | CD1C | CD1c molecule | BDCA1/CD1 | 0.76631 | 0.729005 | 0.447513 | 0.249654 | |||||||||||||||||||||
| B07 | Hs.1799 | NM_001766 | CD1D | CD1d molecule | CD1A/R3 | 0.003685 | 0.002549 | 0.004187 | 0.004095 | |||||||||||||||||||||
| B08 | Hs.523500 | NM_001767 | CD2 | CD2 molecule | SRBC/T11 | 0.003685 | 0.002549 | 0.004187 | 0.004095 | |||||||||||||||||||||
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| # truncated example of Matrix with fold change data (test/control) | |||||||||
| ID_REF | SAMPLE 2/SAMPLE 1 | SAMPLE 3/SAMPLE 1 | SAMPLE 4/SAMPLE 1 | ||||||
| A01 | 0.6916 | 1.136 | 1.1111 | ||||||
| A02 | 1.5454 | 0.9356 | 0.2288 | ||||||
| A03 | 0.6916 | 1.136 | 1.1111 | ||||||
| A04 | 0.6916 | 4.299 | 1.1111 | ||||||
| A05 | 15.327 | 5.4415 | 0.0526 | ||||||
| A06 | 5.056 | 18.5584 | 2.3489 | ||||||
| A07 | 7.3513 | 44.7557 | 1.1111 | ||||||
| A08 | 4.1068 | 25.5278 | 5.3221 | ||||||
| A09 | 8.988 | 133.8061 | 4.7966 | ||||||
| A10 | 2.5456 | 1.136 | 1.1111 | ||||||
| A11 | 240.1846 | 102.1114 | 0.3373 | ||||||
| A12 | 0.8055 | 0.3826 | 0.0215 | ||||||
| B01 | 0.6916 | 1.136 | 1.1111 | ||||||
| B02 | 0.6916 | 1.136 | 1.1111 | ||||||
| B03 | 1.1958 | 0.8259 | 0.026 | ||||||
| B04 | 3.0063 | 3.258 | 1.2501 | ||||||
| B05 | 0.6498 | 0.4305 | 0.274 | ||||||
| B06 | 0.9513 | 0.584 | 0.3258 | ||||||
| B07 | 0.6916 | 1.136 | 1.1111 | ||||||
| B08 | 0.6916 | 1.136 | 1.1111 | ||||||
