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Data Set Group: Hippocampus Consortium M430v2 (Jun06) modify this page

Data Set: Hippocampus Consortium M430v2 (Jun06) PDNN modify this page
GN Accession: GN112
GEO Series: GSE84767
Title: Genetics of the hippocampal transcriptome in mouse: a systematic survey and online neurogenomics resource
Organism: Mouse (mm10)
Group: BXD
Tissue: Hippocampus mRNA
Dataset Status: Public
Platforms: Affy Mouse Genome 430 2.0 (GPL1261)
Normalization: PDNN
Contact Information
Robert Williams
University of Tennessee Health Science Center
855 Monroe Ave Room 501
Memphis, TN 38103 USA
Tel. 901 448-7050
rwilliams@uthsc.edu
Website
Download datasets and supplementary data files

Specifics of this Data Set:
None

Summary:

MOST HIGHLY RECOMMENDED DATA SET (Overall et al., 2009): The Hippocampus Consortium data set provides estimates of mRNA expression in the adult hippocampus of 99 genetically diverse strains of mice including 67 BXD recombinant inbred strains, 13 CXB recombinant inbred strains, a diverse set of common inbred strains, and two reciprocal F1 hybrids.

The hippocampus is an important and intriguing part of the forebrain that is crucial in memory formation and retrieval, and that is often affected in epilepsy, Alzheimer's disease, and schizophrenia. Unlike most other parts of the brain, the hippocampus contains a remarkable population of stems cells that continue to generate neurons and glial cells even in adult mammals (Kempermann, 2005). This genetic analysis of transcript expression in the hippocampus (dentate gyrus, CA1-CA3) is a joint effort of 14 investigators that is supported by numerous agencies described in the acknowledgments section.



About the cases used to generate this set of data:

The BXD genetic reference panel of recombinant inbred strains consists of just over 80 strains. The BXDs in this data set include 27 of the BXD strains made by Benjamin Taylor at the Jackson Laboratory in the 1970s and 1990s (BXD1 through BXD42). All of these strains are fully inbred, many well beyond the 100th filial (F) generation of inbreeding. We have also included 39 inbred (25 strains at F20+) and nearly inbred (14 strains between F14 and F20) BXD lines generated by Lu and Peirce. All of these strains, including those between F14 and F20, have been genotyped at 13,377 SNPs.

Mouse Diversity Panel (MDP). We have profiled a MDP consisting 16 inbred strains and a pair of reciprocal F1 hybrids; B6D2F1 and D2B6F1. These strains were selected for several reasons:

  • genetic and phenotypic diversity, including use by the Phenome Project
  • their use in making genetic reference populations including recombinant inbred strains, cosomic strains, congenic and recombinant congenic strains
  • their use by the Complex Trait Consortium to make the Collaborative Cross (Nairobi/Wellcome, Oak Ridge/DOE, and Perth/UWA)
  • genome sequence data from three sources (NHGRI, Celera, and Perlegen-NIEHS)
  • availability from The Jackson Laboratory

All eight parents of the Collaborative Cross (129, A, C57BL/6J, CAST, NOD, NZO, PWK, and WSB) have been included in the MDP (noted below in the list). Twelve MDP strains have been sequenced, or are currently being resequenced by Perlegen for the NIEHS. This panel will be extremely helpful in systems genetic analysis of a wide variety of traits, and will be a powerful adjunct in fine mapping modulators using what is essentially an association analysis of sequence variants.

  1. 129S1/SvImJ
        Collaborative Cross strain sequenced by NIEHS; background for many knockouts; Phenome Project A list
  2. A/J
        Collaborative Cross strain sequenced by Perlegen/NIEHS; parent of the AXB/BXA panel
  3. AKR/J
        Sequenced by NIEHS; Phenome Project B list
  4. BALB/cByJ
        Sequenced by NIEHS; maternal parent of the CXB panel; Phenome Project A list
  5. BALB/cJ
        Widely used strain with forebrain abnormalities (callosal defects); Phenome Project A list
  6. C3H/HeJ
        Sequenced by Perlegen/NIEHS; paternal parent of the BXH panel; Phenome Project A list
  7. C57BL/6J
        Sequenced by NHGRI; parental strain of AXB/BXA, BXD, and BXH; Phenome Project A list
  8. C57BL/6ByJ
        Paternal substrain of B6 used to generate the CXB panel
  9. CAST/Ei
        Collaborative Cross strain sequenced by NIEHS; Phenome Project A list
  10. DBA/2J
        Sequenced by Perlegen/NIEHS and Celera; paternal parent of the BXD panel; Phenome Project A list
  11. KK/HlJ
        Sequenced by Perlegen/NIEHS
  12. LG/J
        Paternal parent of the LGXSM panel
  13. NOD/LtJ
        Collaborative Cross strain sequenced by NIEHS; Phenome Project B list; diabetic
  14. NZO/HlLtJ
        Collaborative Cross strain
  15. PWD/PhJ
        Sequenced by Perlegen/NIEHS; parental strain for a consomic set by Forjet and colleagues
  16. PWK/PhJ
        Collaborative Cross strain; Phenome Project D list
  17. WSB/EiJ
        Collaborative Cross strain sequenced by NIEHS; Phenome Project C list
  18. B6D2F1 and D2B6F1
    F1 hybrids generated by crossing C57BL/6J with DBA/2J

We have not combined data from reciprocal F1s because they have different Y chromosome and mitochondrial haplotypes. Parent-of-origin effects (imprinting, maternal environment) may also lead to interesting differences in hippocampal transcript levels.

These strains are available from The Jackson Laboratory. BXD43 through BXD100 strains are available from Lu Lu and colleagues at UTHSC.



About the tissue used to generate this set of data:

BXD animals were obtained from UTHSC, UAB, or directly from The Jackson Laboratory (see Table 1 below). Animals were housed at UTHSC, Beth Israel Deaconess, or the Jackson Laboratory before sacrifice. Virtually all CXB animals were obtained directly at the Jackson Laboratory by Lu Lu. We thank Muriel Davisson for making it possible to collect these cases on site. Standard inbred strain stock was from The Jackson Laboratory, but most animals were housed or reared at UTHSC. Mice were killed by cervical dislocation and brains were removed and placed in RNAlater prior to dissection. Cerebella and olfactory bulbs were removed; brains were hemisected, and both hippocampi were dissected whole by Hong Tao Zhang in the Lu lab. Hippocampal samples are very close to complete (see Lu et al., 2001) but probably include variable amounts of subiculum and fimbria.

A great majority of animals used in this study were between 45 and 90 days of age (average of 66 days, maximum range from 41 to 196 days; see Table 1 below). All animals were sacrifice between 9 AM and 5 PM during the light phase.

A pool of dissected tissue typically from six hippocampi and three naive adults of the same strain, sex, and age was collected in one session and used to generate cRNA samples. Two-hundred and one RNA samples were extracted at UTHSC by Zhiping Jia, four samples by Shuhua Qi (R2331H1, R2332H1, P2350H1, R2349H1), and one by Siming Shou (R0129H2).

RNA Extraction: In brief, we used the RNA STAT-60 protocol (TEL-TEST "B" Bulletin No. 1), steps 5.1A (homogenization of tissue), 5.2 (RNA extraction), 5.3 (RNA precipitation), and 5.4 (RNA wash). In Step 5.4 we stopped after adding 75% ethanol (1 ml per 1 ml RNA STAT-60) and stored the mix at -80°C until further use. Before RNA labeling we thawed samples and proceeded with the remainder of Step 5.4; pelleting, drying, and redissolving the pellet in RNase-free water.

We finally purify RNA by using Na4OAc before running arrays. Here is the detailed method:

Final RNA purification protocol

  1. Add 1/10th volume of 3M Na4OAc , pH 5.2. If the sample was eluted with 100 µl nuclease-free water as suggested, this will be 10 µl of 3M Na4OAc.
  2. Add 2.5 volumes of 100% ethanol (250 µl if the RNA was eluted in100 µl). Mix well and incubate at –20°C for 2 hrs.
  3. Centrifuge at speed of 13,000 rpm for 20 min at 4°C. Carefully remove and discard the supernatant.
  4. Wash the pellet with 800 µl 75% cold ethanol, centrifuge at speed of 8,600 rpm for 5 min, and remove the 75% ethanol. Wash again.
  5. To remove the last traces of ethanol, quickly respin the tube, and aspirate any residual fluid.
  6. Air dry the pellet.
  7. Resuspend pellet in nuclease-free water.

 

5. PROTOCOL: RNA/mRNA isolation by the RNA STAT-60 method includes the following steps:
1. Homogenization RNA STAT-60TM (1 ml per 50-100 mg tissue, or 5-10 x 10-6 cells)
2. RNA Extraction 1 vol. of homogenate +0.2 vol. of chloroform
3. RNA Precipitation 0.5 vol. of isopropanol
4. RNA Wash 75% ethanol

Unless stated otherwise the procedure is carried out at room temperature.

5.1 HOMOGENIZATION

A. TISSUES: Homogenize tissues samples in the RNA STAT-60(1 ml/50-100mg tissue) in a glass-Teflon or Polytron homogenizer. Sample volume should not exceed 10% of the volume of the RNA STAT-60 used for homogenization.

B. CELLS: Cells grown in mono layer are lysed directly in a culture dish by adding the RNA STAT-60TM (1 ml/3.5 cm petri dish) and passing the cell lysate several times through a pipette. Cells grown in suspension are sediment then lysed in the RNA STAT-60TM (1 ml per 5-10 x 106 cells) by repetitive pipetting. Washing calls before addition of the RNA STAT-60TM should be avoided as this increases the possibility of mRNA degradation.

5.2 RNA EXTRACTION: Following homogenization, store the homogenate for 5 min at room temp to permit the complete dissociation of nucleoprotein complexes. Next, add 0.2 ml of chloroform per 1 ml of the RNA STAT-60, cover the sample tightly, shake vigorously for 15 seconds and let it stay at room temperature for 2-3minutes. Centrifuge the homogenate at 12,000g (max) for 15 minutes at 4°C. Following centrifugation, the homogenate separates into two phases: a lower red phenol chloroform phase and the colorless upper aqueous phase. RNA remains exclusively in the aqueous phase whereas DNA and proteins are in the interferes and organic phase. The volume of the aqueous phase is about 60% of the volume of RNA STAT-60 used for homogenization.

5.3 RNA PRECIPITATION: Transfer the aqueous phase to a fresh tube and mix with isopropanol. Add 0.5 ml of isopropanol per 1 ml of the RNA STAT-60 used for homogenization. Store samples at room temp for 5-10 minutes and centrifuge at 12,000g (max.) for 10 min at 4°C. RNA precipitate (often visible before centrifugation) forms a white pellet at the bottom of the tube.

5.4 RNA WASH: Remove supernatant and wash the RNA pellet once with 75% ethanol by vortexing and subsequent centrifugation at 7,500g (max.) for 5 min at 4°C. Add at least 1 ml of 75% ethanol per 1 ml of the RNA STAT-60 used for the initial homogenization.

At the end of the procedure, dry the RNA pellet briefly by air-drying or in a vacuum (5-10 min.). It is important not to let the RNA pellet dry completely as it will greatly decrease its solubility. Do not use the Speed-Vac for drying. Dissolve the RNA pellet in water or in 1 mm EDTA, pH 7, or 0.5% SDS solution. Vortex or pass the pellet a few times through a pipette tip. An incubation for 10-15 minutes at 55-60oC may be required to dissolve RNA samples. Diethylpyrocarbonate (DEPC) treated RNase-free solutions1 should be used for solubilization of RNA.

Sample Processing: Samples were processed in the INIA Bioanalytical Core at the W. Harry Feinstone Center for Genomic Research, The University of Memphis, led by Thomas R. Sutter. All processing steps were performed by Shirlean Goodwin. In brief, RNA purity was evaluated using the 260/280 nm absorbance ratio, and values had to be greater than 1.8. The majority of samples were 1.9 to 2.1. RNA integrity was assessed using the Agilent Bioanalyzer 2100. We required an RNA integrity number (RIN) of greater than 8. This RIN value is based on the intensity ratio and amplitude of 18S and 28S rRNA signals. The standard Eberwine T7 polymerase method was used to catalyze the synthesis of cDNA template from polyA-tailed RNA using Superscript II reverse transcriptase (Invitrogen Inc.). The Enzo Life Sciences, Inc., BioArray High Yield RNA Transcript Labeling Kit (T7, Part No. 42655) was used to synthesize labeled cRNA. The cRNA was evaluated using both the 260/280 ratio (values of 2.0 or 2.1 are acceptable) and the Bioanalyzer output (a dark cRNA smear on the 2100 output centered roughly between 600 and 2000 nucleotides is required). Those samples that passed both QC steps (10% usually failed and new RNA samples had to be acquired and processed) were then sheared using a fragmentation buffer included in the Affymetrix GeneChip Sample Cleanup Module (Part No. 900371). Fragmented cRNA samples were either stored at -80°C until use or were immediately injected onto the array. The arrays were hybridized and washed following standard Affymetrix protocols.

Replication and Sample Balance: Our goal was to obtain a male sample pool and female sample pool from each isogenic group. While almost all strains were orginally represented by matched male and female samples, not all data sets passed the final quality control steps. All but 5 of 99 strains (BXD55, BXD86, BXD94, BALB/cByJ, and CAST/EiJ) are represented by pairs or (rarely) trios of arrays. The first and last samples are technical replicates of a B6D2F1 hippocampal pool (aliquots R1291H3 and R1291H4).

 

Sex Balance: Based on the expression of Xist, probe set 1427262_at, DBA/2J and KK/HlJ are represented only by female samples, BXD55, and BALB/cByJ are only represented by a single male sample, BXD74 is represented by two male samples, and BXD86, BXD94, and CAST/EiJ are possibly mixed sex samples. One of the BXD9 samples, array R1523, may be a mixed sex sample pool because the expression of Xist is intermediate.

Experimental Design and Batch Structure: This data set consists arrays processed in six groups over a three month period (May 2005 to August 2005). Each group consists of 32 to 34 arrays. Sex, strain, and strain type (BXD, CXB, and MDP) were interleaved among groups to ensure reasonable balance and to minimize group-by-strain statistical confounds in group normalization. The two independent samples from a single strain were always run in different groups. All arrays were processed using a single protocol by a single operator, Shirlean Goodwin.

All samples in a group were labeled on one day, except for a few cases that failed QC on their first pass. The hybridization station accommodates up to 20 samples, and for this reason each group was split into a large first set of 20 samples and a second set of 12 to 14 samples. Samples were washed in groups of four and then held in at 4°C until all 20 (or 12-14) arrays were ready to scan. The last four samples out of the wash stations were scanned directly. Samples were scanned in sets of four.

COMPARISON with December 2005 Data Set: Both BXD14 arrays in the Dec05 data set were found to actually be from BXD23 cases. This error of strain identification has been corrected in the present data set. Four arrays in the Dec05 data set have been deleted because we judged them to be of poor quality (strain_sex_sample_firstreaction_group):

  1. BXD21_F_1_1_G1
  2. BXD23_M_1_1_G7
  3. BXD36_M_1_1_G2
  4. BXD36_F_1_1_G3

In the Dec05 data set there are a total of 1986 transcripts with QTLs that have LRS scores above 50, whereas in the corrected June06 data sets there are a total of 2074 transcripts with QTLs above 50.

Data Table 1:

This table lists all arrays by file order (Index), tube/sample ID, age, sex, batch, and numbers of animals in each sample pool (pool size). The next columns (RMA outlier, scale factor, background average, present, absent, marginal, AFFY-b-ActinMur(3'/5'), AFFY-GapdhMur(3'/5')) are all Affymetrix QC data. Finally, source lists the source colony of the animals. (Final version, fully corrected, by Arthur Centeno, October 2008)

index tube ID strain age sex batch ID pool size RMA outlier scale factor back ground average present absent marginal AFFX-b-ActinMur (3'/5') AFFX-GapdhMur (3'/5') source
1 R1289H2 B6D2F1 64 F 6 3 0.02 2.406 53.84 0.492 0.489 0.019 1.61 0.96 UTM RW
2 R1291H3 B6D2F1 66 M 1 3 0.01 3.524 48.54 0.487 0.494 0.019 1.21 1.52 UTM RW
3 R1291H4 B6D2F1 66 M technical duplicate of above 6 3 0.08 3.891 46.69 0.512 0.469 0.019 1.9 0.89 UTM RW
4 R2045H2 D2B6F1 65 F 1 2 0.01 4.403 47.99 0.497 0.485 0.018 1.09 1.53 UTM RW
5 R1595H2 D2B6F1 63 F 5 3 0.06 2.579 58.49 0.506 0.475 0.019 2.49 1.21 UTM RW
6 R1551H1 D2B6F1 72 F 6 3 0.02 2.62 53.76 0.506 0.476 0.018 1.37 0.76 UTM RW
7 R1361H1 C57BL/6J 69 F 6 4 0.01 3.058 51.87 0.477 0.503 0.02 1.67 0.76 UTM RW
8 R2041H2 C57BL/6J 65 M 1 4 0.04 3.341 49.26 0.527 0.456 0.018 1.14 1.45 UTM RW
9 R1449H2 C57BL/6J 71 M 5 3 0.09 3.592 44.32 0.47 0.51 0.02 1.68 0.77 UTM DG
10 R1290H2 DBA/2J 63 F 7 2 0.04 2.576 59.6 0.513 0.468 0.018 1.3 0.78 JAX
11 R1468H1 DBA/2J 64 F 5 3 0.03 2.929 53.8 0.515 0.465 0.019 1.28 0.79 UTM RW
12 R1507H1 BXD1 58 M 3 3 0.02 4.056 60.17 0.478 0.503 0.019 1.15 0.76 Glenn
13 R1542H1 BXD1 59 F 7 3 0.03 1.792 80.56 0.492 0.489 0.018 1.57 0.79 Glenn
14 R1520H1 BXD2 56 F 4 4 0.09 1.715 71.62 0.515 0.467 0.018 2.36 1.6 Glenn
15 R1516H1 BXD2 61 M 1 4 0.01 2.231 64.86 0.508 0.474 0.019 1.3 1.53 Glenn
16 R1593H2 BXD5 60 F 1 4 0 1.913 59.96 0.487 0.493 0.02 0.98 1.44 Glenn
17 R1692H1 BXD5 60 M 3 2 0.07 3.764 72.74 0.465 0.516 0.02 1.15 0.74 Glenn
18 R1539H2 BXD6 59 F 1 4 0 2.488 54.97 0.518 0.463 0.018 1.08 1.33 Glenn
19 R1538H1 BXD6 59 M 4 3 0.01 2.585 50.27 0.505 0.475 0.02 1.46 0.79 Glenn
20 R1518H1 BXD8 56 F 1 3 0 2.92 54.84 0.515 0.465 0.02 1.32 1.24 Glenn
21 R1548H1 BXD8 59 M 6 3 0.07 2.132 59.37 0.504 0.477 0.019 2.16 1.54 Glenn
22 R1350H2 BXD9 86 F 1 3 0.05 2.771 60.62 0.5 0.482 0.018 1.01 1.28 UMemphis
23 R1523H3 BXD9 57 MF (mixed) 7 3 0.14 3.9 78.36 0.435 0.547 0.018 1.36 0.77 UTM RW
24 R1531H1 BXD11 56 F 6 3 0.06 2.229 56.36 0.505 0.475 0.02 2.23 1.02 Glenn
25 R1367H1 BXD11 56 M 1 3 0.01 2.11 78.78 0.503 0.477 0.02 1.07 1.27 Glenn
26 R1530H1 BXD12 58 F 1 3 0 3.227 53.77 0.505 0.477 0.018 0.95 1.4 Glenn
27 R2674H1 BXD12 59 M 7 3 0.03 1.924 83.44 0.519 0.464 0.018 1.21 0.78 Glenn
28 R1529H1 BXD13 58 F 6 3 0.05 2.55 59.05 0.497 0.485 0.018 2 1.54 Glenn
29 R1662H2 BXD13 60 M 1 3 0.03 4.603 45.81 0.509 0.472 0.019 1.3 0.82 Glenn
30 R1304H2 BXD14 72 F 7 3 0.03 3.946 61.87 0.484 0.498 0.018 1.22 0.77 UTM RW
31 R1278H2 BXD14 55 M 7 3 0.06 4.75 67.52 0.449 0.532 0.019 1.1 0.73 UTM RW
32 R1524H1 BXD15 60 F 6 4 0.02 2.961 50.93 0.497 0.484 0.019 1.74 0.91 Glenn
33 R1515H1 BXD15 61 M 1 3 0.01 3.316 57.05 0.503 0.478 0.019 1.32 1.21 Glenn
34 R1661H1 BXD16 61 F 1 3 0.01 2.778 59.81 0.516 0.466 0.019 1.39 1.2 Glenn
35 R1594H1 BXD16 61 M 4 3 0.03 2.634 53.66 0.504 0.478 0.018 1.96 1.51 Glenn
36 R2666H1 BXD19 60 F 7 3 0.02 2.498 76.2 0.495 0.486 0.019 1.41 0.77 Glenn
37 R1471H1 BXD19 157 M 1 3 0.02 3.165 43.34 0.519 0.462 0.018 1.01 1.29 UTM JB
38 R1573H1 BXD20 59 F 1 3 0.02 3.749 52.7 0.513 0.469 0.018 1.01 1.27 Glenn
39 R2507H1 BXD20 60 M 6 3 0.06 3.568 57 0.472 0.508 0.02 1.29 0.76 Glenn
40 R2668H1 BXD21 60 M 7 4 0.07 2.605 44.9 0.535 0.449 0.017 1.54 0.76 Glenn
41 R1337H2 BXD21 102 F 2 4 0 2.673 58.05 0.492 0.489 0.019 1.4 0.76 UAB
42 R1848H3 BXD22 196 F 6 4 0.02 2.943 51.7 0.494 0.485 0.021 2.2 0.78 UAB
43 R1525H1 BXD22 59 M 2 3 0.02 2.248 55.76 0.548 0.433 0.018 1.26 0.74 Glenn
44 R1280H2 BXD23 56 F 1 3 0.01 3.187 54.63 0.458 0.523 0.019 0.96 1.2 UTM RW
45 R1537H1 BXD23 58 F 5 3 0.1 3.719 67.54 0.468 0.513 0.019 1.51 0.96 Glenn
46 R1343H2 BXD24 71 F 2 3 0.01 2.083 65.07 0.506 0.474 0.019 1.46 0.75 UMemphis
47 R1517H1 BXD24 57 M 3 3 0.01 3.471 53.66 0.504 0.476 0.019 1.28 0.78 Glenn
48 R1366H1 BXD27 60 F 2 4 0 2.26 48.46 0.518 0.463 0.019 1.29 0.77 Glenn
49 R1849H1 BXD27 70 M 5 3 0.06 8.801 38.34 0.468 0.512 0.019 2.42 1.08 UAB
50 R1353H1 BXD28 79 F 3 4 0.01 3.22 76.22 0.48 0.5 0.02 1.33 0.78 UMemphis
51 R2332H1 BXD28 60 M 2 3 0.01 3.217 63.68 0.491 0.49 0.019 1.37 0.79 Glenn
52 R1532H1 BXD29 57 F 2 3 0.01 2.122 59.18 0.524 0.456 0.019 1.17 0.76 Glenn
53 R1356H1 BXD29 76 M 5 3 0.01 4.033 47.67 0.52 0.463 0.017 1.17 0.78 UMemphis
54 R1240H2 BXD31 61 M 2 3 0.02 2.335 65.17 0.507 0.474 0.019 1.31 0.78 UTM RW
55 R1526H2 BXD31 57 F 7 4 0.1 7.267 89.54 0.435 0.547 0.017 1.35 0.78 UTM RW
56 R2675H1 BXD32 57 F 7 3 0.03 2.268 78.01 0.502 0.478 0.02 1.22 0.78 Glenn
57 R1508H2 BXD32 58 M 2 4 0.01 1.917 67.78 0.539 0.442 0.019 1.28 0.73 Glenn
58 R1345H3 BXD33 65 F 2 2 0.01 2.098 63.14 0.522 0.459 0.019 1.27 0.73 UMemphis
59 R1581H1 BXD33 59 M 3 3 0.01 3.229 53.16 0.496 0.485 0.019 1.19 0.78 Glenn
60 R1527H1 BXD34 59 F 2 3 0.01 2.3 58.92 0.51 0.471 0.019 1.24 0.76 Glenn
61 R1339H3 BXD34 74 M 5 3 0.12 2.888 53.49 0.506 0.476 0.018 2.39 1.35 UMemphis
62 R1855H1 BXD38 55 F 3 4 0.01 3.536 54.54 0.49 0.492 0.018 1.39 0.75 Glenn
63 R1510H1 BXD38 59 M 2 3 0.01 2.186 68.06 0.521 0.46 0.019 1.26 0.79 Glenn
64 R1528H2 BXD39 59 F 2 3 0.03 4.717 38.3 0.511 0.47 0.02 1.12 0.75 Glenn
65 R1514H1 BXD39 59 M 3 3 0.03 3.992 56.06 0.477 0.504 0.019 1.43 0.81 Glenn
66 R1522H1 BXD40 59 F 4 4 0 2.631 67.16 0.49 0.491 0.018 1.56 0.77 Glenn
67 R1359H1 BXD40 73 M 2 3 0.09 7.458 39.86 0.451 0.527 0.021 1.28 0.74 UMemphis
68 R1541H2 BXD42 58 F 7 3 0.07 6.784 52.12 0.483 0.499 0.017 1.13 0.66 Glenn
69 R1540H1 BXD42 58 M 7 4 0.03 2.423 75.14 0.492 0.488 0.02 1.48 0.78 Glenn
70 R1334H2 BXD43 59 F 1 3 0 2.672 54.36 0.492 0.491 0.017 1.2 2.06 UTM RW
71 R1303H1 BXD43 63 M 3 4 0.02 3.497 51.9 0.486 0.495 0.019 1.15 0.8 UTM RW
72 R1326H1 BXD44 65 F 3 4 0 3.412 53.96 0.496 0.485 0.018 1.35 0.78 UTM RW
73 R1577H2 BXD44 56 M 1 3 0.02 2.159 67.52 0.512 0.469 0.019 1.18 1.71 UTM RW
74 R1403H2 BXD45 63 F 7 2 0.03 3.146 44.5 0.524 0.457 0.018 1.41 0.78 Glenn
75 R1472H1 BXD45 65 M 7 4 0.04 1.651 73.31 0.543 0.44 0.018 1.63 0.74 UTM RW
76 R1316H1 BXD48 58 F 4 3 0 2.445 68.59 0.515 0.467 0.019 1.16 0.73 UTM RW
77 R1575H3 BXD48 65 M 3 4 0.05 4.577 55.78 0.466 0.514 0.019 1.59 0.9 UTM RW
78 R2521H1 BXD50 63 F 6 4 0.01 3.109 57.28 0.495 0.485 0.02 1.23 0.78 UTM RW
79 R1944H2 BXD50 81 M 1 3 0.01 2.546 63.39 0.495 0.485 0.02 0.9 1.57 UTM RW
80 R2331H1 BXD51 66 F 3 3 0.03 3.534 44.42 0.501 0.481 0.017 1.2 0.9 UTM RW
81 R1582H1 BXD51 71 M 6 4 0.03 2.92 47.87 0.489 0.491 0.02 1.36 0.75 UTM RW
82 R2680H1 BXD55 65 M 7 3 0.07 1.707 79.75 0.503 0.48 0.017 1.91 1.05 UTM RW
83 R1331H1 BXD60 60 F 4 3 0.01 2.867 50.33 0.492 0.487 0.021 1.34 0.78 UTM RW
84 R1281H2 BXD60 59 M 1 3 0 2.39 58.44 0.511 0.469 0.02 0.94 1.2 UTM RW
85 R2667H1 BXD61 70 F 7 4 0.03 3.36 59.04 0.495 0.488 0.018 1.16 0.76 UTM RW
86 R1856H2 BXD61 94 M 1 2 0 3.502 49.6 0.501 0.48 0.019 0.96 1.3 UTM RW
87 R1246H1 BXD62 54 F 1 4 0.02 3.405 51.47 0.511 0.471 0.018 1.14 1.34 UTM RW
88 R1585H2 BXD62 64 M 6 4 0.01 3.156 55.77 0.518 0.464 0.018 1.43 0.82 UTM RW
89 R1945H1 BXD63 107 F 1 3 0.02 2.811 52.65 0.522 0.459 0.019 1.05 1.36 UTM RW
90 R2093H3 BXD63 70 M 6 3 0.02 3.894 42.85 0.503 0.477 0.019 1.29 1.01 UTM RW
91 R2062H2 BXD64 65 F 1 3 0.05 3.795 78.48 0.513 0.468 0.019 0.98 1.43 UTM RW
92 R2061H1 BXD64 87 M 3 4 0.01 3.536 61.57 0.477 0.504 0.019 1.31 0.78 UTM RW
93 R2054H2 BXD65 55 F 1 2 0.03 3.159 80.96 0.48 0.502 0.018 1.09 1.24 UTM RW
94 R2056H2 BXD65 89 M 6 2 0 2.836 59.6 0.504 0.477 0.019 1.3 0.75 UTM RW
95 R1941H2 BXD66 78 F 1 4 0.01 2.734 50.93 0.499 0.481 0.02 1.18 1.29 UTM RW
96 R1949H2 BXD66 96 M 4 2 0.04 2.828 51.27 0.474 0.508 0.019 2.05 1.12 UTM RW
97 R2060H1 BXD67 54 F 6 3 0.01 2.561 43.88 0.502 0.479 0.02 1.7 0.84 UTM RW
98 R2052H1 BXD67 61 M 1 4 0.01 3.161 43.23 0.521 0.46 0.018 1.09 1.31 UTM RW
99 R2074H1 BXD68 60 F 5 3 0.02 6.528 49.62 0.479 0.502 0.019 1.48 0.83 UTM RW
100 R1928H1 BXD68 72 M 2 2 0.01 2.404 48.28 0.521 0.459 0.02 1.3 0.74 UTM RW
101 R1439H3 BXD69 60 F 2 3 0.02 2.463 59.14 0.522 0.459 0.018 1.31 0.78 UTM RW
102 R1559H1 BXD69 64 M 3 3 0.03 2.987 67.74 0.486 0.496 0.017 1.38 0.8 UTM RW
103 R2134H1 BXD70 64 F 5 2 0.02 2.148 58.64 0.532 0.45 0.019 1.4 0.85 UTM RW
104 R2063H1 BXD70 55 M 2 3 0.02 3.481 55.32 0.513 0.469 0.018 1.28 0.71 UTM RW
105 R1277H1 BXD73 60 F 4 2 0.01 2.576 62.45 0.502 0.479 0.019 1.35 0.79 UTM RW
106 R1443H2 BXD73 76 M 2 3 0.01 2.312 64.34 0.499 0.481 0.02 1.48 0.77 UTM RW
107 R2055H2 BXD74 79 M 2 3 0.01 2.576 56.84 0.509 0.473 0.018 1.46 0.88 UTM RW
108 R2316H1 BXD74 193 M 5 2 0.01 3.457 55.35 0.508 0.471 0.02 1.17 0.78 UTM RW
109 R1871H1 BXD75 61 F 2 3 0.04 1.723 56.4 0.53 0.451 0.019 1.3 0.76 UTM RW
110 R1844H2 BXD75 90 M 3 4 0.01 1.934 56.23 0.52 0.461 0.019 1.62 0.86 UTM RW
111 R1948H2 BXD76 81 F 2 3 0.01 1.507 68.85 0.553 0.428 0.02 1.3 0.75 UTM RW
112 R2094H1 BXD76 61 M 5 4 0.01 3.299 42.69 0.519 0.462 0.019 1.39 0.88 UTM RW
113 R2262H1 BXD77 62 F 3 4 0.02 4.317 47.16 0.493 0.488 0.019 1.32 0.74 UTM RW
114 R1423H1 BXD77 62 M 2 3 0.02 3.071 54.15 0.51 0.471 0.019 1.26 0.74 UTM RW
115 R1947H1 BXD79 108 F 2 2 0.01 2.599 51.52 0.524 0.457 0.019 1.35 0.74 UTM RW
116 R2092H1 BXD79 86 M 5 4 0.06 3.735 42.25 0.514 0.468 0.018 2.94 1.06 UTM RW
117 R1880H1 BXD80 68 F 5 3 0.06 4.855 42.22 0.501 0.481 0.018 2.17 1.36 UTM RW
118 R1881H2 BXD80 68 M 2 3 0.02 2.073 48.93 0.524 0.458 0.019 1.34 0.83 UTM RW
119 R2075H1 BXD83 60 F 2 3 0.01 2.454 55.1 0.502 0.48 0.018 1.27 0.77 UTM RW
120 R2076H2 BXD83 60 M 6 3 0.03 2.624 55.65 0.495 0.488 0.018 2.21 0.94 UTM RW
121 R2077H2 BXD84 62 F 6 2 0 2.1 71.87 0.522 0.459 0.018 1.68 0.81 UTM RW
122 R2135H3 BXD84 75 M 2 2 0.01 2.467 64.46 0.505 0.476 0.019 1.2 0.74 UTM RW
123 R1473H1 BXD85 79 F 2 3 0.02 3.384 55.34 0.478 0.502 0.02 1.24 0.77 UTM RW
124 R1474H1 BXD85 57 M 1 3 0.01 2.831 55.24 0.522 0.461 0.018 1.04 1.29 UTM RW
125 R1597H1 BXD85 86 M 4 4 0.09 2.028 53.95 0.487 0.492 0.021 1.28 0.83 UTM RW
126 R1415H1 BXD86 77 F 4 3 0.02 2.525 53.16 0.495 0.485 0.02 1.66 0.91 UTM RW
127 R2669H2 BXD87 63 F 7 3 0.07 2.61 57.59 0.513 0.47 0.018 1.6 0.91 UTM RW
128 R1710H1 BXD87 84 M 2 4 0.01 2.697 56.4 0.512 0.469 0.019 1.28 0.79 UTM RW
129 R1872H2 BXD89 90 F 2 2 0.02 3.013 63.53 0.492 0.488 0.021 1.22 0.72 UTM RW
130 R1850H3 BXD89 82 M 4 4 0.03 2.736 44.89 0.498 0.483 0.019 1.5 0.83 UTM RW
131 R2058H1 BXD90 61 F 2 3 0.01 3.389 48.05 0.502 0.478 0.02 1.53 0.76 UTM RW
132 R1600H2 BXD90 74 M 7 4 0.03 3.261 51.31 0.517 0.465 0.018 1.16 0.75 Glenn
133 R1301H2 BXD92 58 F 2 3 0.02 3.543 41.97 0.522 0.46 0.018 1.5 0.79 UTM RW
134 R1309H1 BXD92 59 M 4 3 0.05 1.655 66.34 0.498 0.481 0.021 1.52 0.82 UTM RW
135 R2057H1 BXD93 92 F 5 3 0.02 4.033 44.41 0.509 0.471 0.02 1.22 0.78 UTM RW
136 R2059H1 BXD93 58 M 1 3 0 3.058 60.29 0.493 0.488 0.019 1.18 1.37 UTM RW
137 R2313H1 BXD94 59 F 3 3 0 3.091 59.45 0.487 0.495 0.018 1.34 0.73 UTM RW
138 R1915H1 BXD96 65 F 5 2 0.04 5.145 46.19 0.502 0.481 0.017 1.37 0.74 UTM RW
139 R1846H2 BXD96 63 M 1 3 0 3.159 55.85 0.487 0.493 0.02 0.92 1.26 UTM RW
140 R2648H1 BXD97 74 F 7 4 0.02 1.664 82.08 0.518 0.464 0.019 1.4 0.78 UTM RW
141 R1927H2 BXD97 67 M 1 3 0.04 2.622 57.81 0.539 0.444 0.017 1.45 1.32 UTM RW
142 R1942H1 BXD98 62 F 5 3 0.04 3.104 48.42 0.528 0.454 0.019 2.22 1.08 UTM RW
143 R1943H2 BXD98 62 M 3 3 0.02 4.04 56.85 0.484 0.497 0.019 1.18 0.76 UTM RW
144 R2197H1 BXD99 70 F 3 3 0.02 4.288 51.75 0.49 0.492 0.018 1.35 0.81 UTM RW
145 R2315H1 BXD99 84 M 5 2 0.03 6.036 43.05 0.484 0.497 0.018 1.7 0.96 UTM RW
146 R2028H2 129S1/SvImJ 66 F 5 3 0.1 4.362 64.49 0.497 0.484 0.019 2.78 1.13 JAX
147 R2029H2 129S1/SvImJ 66 M 6 3 0.04 5.208 41.21 0.49 0.49 0.02 1.62 0.95 JAX
148 R2670H1 A/J 65 F 7 3 0.04 3.951 46.8 0.498 0.485 0.017 1.32 0.75 UTM RW
149 R2030H1 A/J 57 M 5 2 0.06 3.307 45.16 0.527 0.454 0.018 1.63 0.99 UTM RW
150 R2032H3 AKR/J 66 F 5 3 0.04 3.054 61.03 0.51 0.471 0.018 1.46 0.79 JAX
151 R2454H1 AKR/J 66 M 6 4 0.11 2.892 58.55 0.474 0.507 0.019 1.99 0.78 JAX
152 R1675H1 BALB/cByJ 83 M 7 3 0.03 3.405 48.13 0.509 0.474 0.018 1.13 0.78 JAX
153 R2036H3 BALB/cJ 51 F 5 3 0.12 2.611 56.29 0.518 0.466 0.017 3.3 1.23 UTM RW
154 R2053H1 BALB/cJ 55 M 5 3 0.1 2.505 63.27 0.499 0.483 0.018 3.1 1.34 UTM RW
155 R2037H2 BALB/cJ 51 M 6 4 0.01 2.546 58.13 0.497 0.485 0.018 1.26 0.77 UTM RW
156 R2038H3 C3H/HeJ 63 F 6 3 0.02 2.671 66.74 0.476 0.504 0.02 1.41 0.77 UTM RW
157 R2039H1 C3H/HeJ 63 M 5 3 0.1 3.384 44.15 0.528 0.454 0.017 2.16 0.88 UTM RW
158 R2137H1 C57BL/6ByJ 55 F 5 3 0.02 4.746 47.01 0.488 0.493 0.018 1.23 0.79 JAX
159 R2673H1 C57BL/6ByJ 55 M 7 3 0.08 1.842 67.69 0.514 0.469 0.017 1.75 0.78 JAX
160 R2619H1 CAST/EiJ 64 F 5 3 0.14 4.077 51.87 0.455 0.528 0.018 2.74 1.2 JAX
161 R1683H1 KK/HIJ 72 F 6 3 0.02 3.919 54.23 0.491 0.489 0.02 1.31 0.83 JAX
162 R1687H3 KK/HIJ 72 F 5 3 0.04 3.888 40.86 0.499 0.483 0.019 1.86 0.88 JAX
163 R2046H1 LG/J 63 F 5 2 0.03 2.822 59.18 0.514 0.468 0.018 1.68 0.8 UTM RW
164 R2047H2 LG/J 63 M 6 3 0.07 2.038 60.34 0.509 0.471 0.02 2.16 0.95 UTM RW
165 R2048H1 NOD/LtJ 77 F 6 2 0.14 4.045 50.21 0.489 0.49 0.021 2.89 0.95 UTM RW
166 R2049H3 NOD/LtJ 76 M 5 3 0.1 2.328 52.78 0.519 0.462 0.019 3.09 1.35 UTM RW
167 R2200H1 NZO/HlLtJ 62 F 5 2 0.03 2.648 54.29 0.543 0.438 0.019 1.27 0.8 JAX
168 R2350H1 NZO/HlLtJ 96 M 6 2 0.19 2.391 50.52 0.518 0.463 0.02 3.71 2.21 JAX
169 R2677H1 PWD/PhJ 65 M 7 2 0.12 2.764 65.49 0.462 0.52 0.018 1.89 1.16 UTM RW
170 R2051H3 PWD/PhJ 64 M 5 3 0.07 3.266 51.5 0.475 0.506 0.019 2.8 1.01 UTM RW
171 R2322H1 PWK/PhJ 63 F 5 2 0.09 2.94 54.91 0.511 0.47 0.019 2.32 1.02 JAX
172 R2349H1 PWK/PhJ 83 M 6 2 0.15 3.306 54.93 0.459 0.522 0.019 4.65 1.45 JAX
173 R2198H2 WSB/EiJ 58 F 6 1 0.02 2.922 57.97 0.502 0.479 0.019 1.44 0.76 JAX
174 R2199H1 WSB/EiJ 58 M 5 3 0.04 3.171 54.95 0.475 0.505 0.02 1.32 0.81 JAX
175 R2116H1 CXB1 55 F 3 3 0.07 5.792 51.59 0.459 0.521 0.02 1.17 0.8 JAX
176 R2096H1 CXB1 55 M 4 2 0.01 3.435 53.78 0.495 0.485 0.02 1.22 0.79 JAX
177 R2117H2 CXB2 62 F 4 2 0.04 3.39 45.97 0.533 0.45 0.017 2.05 0.89 JAX
178 R2098H1 CXB2 68 M 3 3 0.02 2.572 54.22 0.496 0.485 0.019 1.38 0.86 JAX
179 R2118H1 CXB3 47 F 3 3 0.03 3.646 63.16 0.478 0.503 0.019 1.22 0.77 JAX
180 R2100H1 CXB3 47 M 4 3 0.02 5.76 51.38 0.48 0.503 0.017 1.24 0.81 JAX
181 R2119H1 CXB4 58 F 4 3 0.02 3.897 49.21 0.488 0.494 0.018 1.31 0.79 JAX
182 R2101H1 CXB4 58 M 3 3 0.13 7.372 53.77 0.433 0.548 0.019 1.2 0.97 JAX
183 R2505H1 CXB5 80 F 6 3 0.02 2.83 49.6 0.499 0.48 0.02 1.33 0.76 UTM RW
184 R2131H1 CXB5 42 M 4 3 0.1 5.577 51.15 0.434 0.547 0.019 1.7 0.89 JAX
185 R0129H2 CXB5 70 M 3 3 0.07 4.829 45.42 0.488 0.493 0.019 1.23 0.83 UTM RW
186 R2676H1 CXB6 47 F 7 2 0.05 2.146 62.51 0.507 0.475 0.018 1.52 0.78 JAX
187 R2102H1 CXB6 49 M 4 3 0.07 5.148 51.63 0.453 0.529 0.018 1.43 0.87 JAX
188 R2121H1 CXB7 63 F 4 2 0.06 4.904 48.71 0.464 0.517 0.019 1.19 0.92 JAX
189 R2104H2 CXB7 58 M 3 2 0.06 3.389 48.79 0.502 0.479 0.019 1.74 1.48 JAX
190 R2122H1 CXB8 54 F 3 3 0.04 4.128 59.77 0.451 0.529 0.02 1.12 0.76 JAX
191 R2105H1 CXB8 41 M 4 3 0.16 3.146 61.04 0.451 0.53 0.019 1.34 0.84 JAX
192 R2123H1 CXB9 54 F 3 3 0.08 5.708 55.94 0.438 0.543 0.019 1.32 0.78 JAX
193 R2106H1 CXB9 54 M 4 3 0.06 5.868 46.55 0.469 0.512 0.019 1.18 0.82 JAX
194 R2124H1 CXB10 53 F 4 2 0.11 4.867 39.88 0.451 0.528 0.02 1.55 0.8 JAX
195 R2671H1 CXB10 53 M 7 3 0.09 2.348 71.45 0.488 0.494 0.018 2.2 1.14 JAX
196 R2125H1 CXB11 58 F 3 3 0.03 3.256 54.95 0.461 0.519 0.02 1.46 0.77 JAX
197 R2128H1 CXB11 58 M 4 2 0.06 4.986 54.13 0.465 0.515 0.02 1.11 0.83 JAX
198 R2126H1 CXB12 47 F 4 3 0.11 3.935 54.11 0.469 0.511 0.021 1.5 0.79 JAX
199 R2109H1 CXB12 47 M 3 3 0.07 4.518 49.26 0.488 0.492 0.02 1.23 0.77 JAX
200 R2672H1 CXB13 49 F 7 3 0.03 1.722 79.52 0.516 0.465 0.019 1.64 0.75 JAX
201 R2110H1 CXB13 56 M 4 3 0.21 3.478 48.08 0.461 0.517 0.022 1.21 0.78 JAX


About the array platform:

Affymetrix Mouse Genome 430 2.0 array: The 430v2 array consists of 992936 useful 25-nucleotide probes that estimate the expression of approximately 39,000 transcripts and the majority of known genes and expressed sequence tags. The array sequences were selected late in 2002 using Unigene Build 107 by Affymetrix. The UTHSC group has recently reannotated all probe sets on this array, producing more accurate data on probe and probe set targets. All probes were aligned to the most recent assembly of the Mouse Genome (Build 34, mm6) using Jim Kent's BLAT program. Many of the probe sets have been manually curated by Jing Gu and Rob Williams.



About data values and data processing:

Harshlight was used to examine the image quality of the array (CEL files). Bad areas (bubbles, scratches, blemishes) of arrays were masked.

First pass data quality control: Affymetrix GCOS provides useful array quality control data including:

  1. The scale factor used to normalize mean probe intensity. This averaged 3.3 for the 179 arrays that passed and 6.2 for arrays that were excluded. The scale factor is not a particular critical parameter.
  2. The average background level. Values averaged 54.8 units for the data sets that passed and 55.8 for data sets that were excluded. This factor is not important for quality control.
  3. The percentage of probe sets that are associated with good signal ("present" calls). This averaged 50% for the 179 data sets that passed and 42% for those that failed. Values for passing data sets extended from 43% to 55%. This is a particularly important criterion.
  4. The 3':5' signal ratios of actin and Gapdh. Values for passing data sets averaged 1.5 for actin and 1.0 for Gapdh. Values for excluded data sets averaged 12.9 for actin and 9.6 for Gapdh. This is a highly discriminative QC criterion, although one must keep in mind that only two transcripts are being tested. Sequence variation among strains (particularly wild derivative strains such as CAST/Ei) may affect these ratios.

The second step in our post-processing QC involves a count of the number of probe sets in each array that are more than 2 standard deviations (z score units) from the mean across the entire 206 array data sets. This was the most important criterion used to eliminate "bad" data sets. All 206 arrays were processed togther using standard RMA and PDNN methods. The count and percentage of probe sets in each array that were beyond the 2 z theshold was computed. Using the RMA transform the average percentage of probe sets beyond the 2 z threshold for the 179 arrays that finally passed of QC procedure was 1.76% (median of 1.18%). In contrast the 2 z percentage was more than 10-fold higher (mean of 22.4% and median 20.2%) for those arrays that were excluded. This method is not very sensitive to the transformation method that is used. Using the PDNN transform, the average percent of probe sets exceeding was 1.31% for good arrays and was 22.6% for those that were excluded. In our opinion, this 2 z criterion is the most useful criterion for the final decision of whether or not to include arrays, although again, allowances need to be made for wild strains that one expects to be different from the majority of conventional inbred strains. For example, if a data set has excellent characteristics on all of the Affymetrix GCOS metrics listed above, but generates a high 2 z percentage, then one would include the sample if one can verify that there are no problems in sample and data set identification.

The entire procedure can be reapplied once the initial outlier data sets have been eliminated to detect any remaining outlier data sets.

DataDesk was used to examine the statistical quality of the probe level (CEL) data after step 5 below. DataDesk allows the rapid detection of subsets of probes that are particularly sensitive to still unknown factors in array processing. Arrays can then be categorized at the probe level into "reaction classes." A reaction class is a group of arrays for which the expression of essentially all probes are colinear over the full range of log2 values. A single but large group of arrays (n = 32) processed in essentially the identical manner by a single operator can produce arrays belonging to as many as four different reaction classes. Reaction classes are NOT related to strain, age, sex, treatment, or any known biological parameter (technical replicates can belong to different reaction classes). We do not yet understand the technical origins of reaction classes. The number of probes that contribute to the definition of reaction classes is quite small (<10% of all probes). We have categorized all arrays in this data set into one of 5 reaction classes. These have then been treated as if they were separate batches. Probes in these data type "batches" have been aligned to a common mean as described below.

Probe (cell) level data from the CEL file: These CEL values produced by GCOS are 75% quantiles from a set of 91 pixel values per cell.

  1. We added an offset of 1.0 unit to each cell signal to ensure that all values could be logged without generating negative values. We then computed the log base 2 of each cell.
  2. We performed a quantile normalization of the log base 2 values for all arrays using the same initial steps used by the RMA transform.
  3. We computed the Z scores for each cell value.
  4. We multiplied all Z scores by 2.
  5. We added 8 to the value of all Z scores. The consequence of this simple set of transformations is to produce a set of Z scores that have a mean of 8, a variance of 4, and a standard deviation of 2. The advantage of this modified Z score is that a two-fold difference in expression level (probe brightness level) corresponds approximately to a 1 unit difference.
  6. Finally, we computed the arithmetic mean of the values for the set of microarrays for each strain. Technical replicates were averaged before computing the mean for independent biological samples. Note, that we have not (yet) corrected for variance introduced by differences in sex or any interaction terms. We have not corrected for background beyond the background correction implemented by Affymetrix in generating the CEL file. We eventually hope to add statistical controls and adjustments for some of these variables.

Probe set data from the CHP file: The expression values were generated using PDNN. The same simple steps described above were also applied to these values. Every microarray data set therefore has a mean expression of 8 with a standard deviation of 2. A 1 unit difference represents roughly a two-fold difference in expression level. Expression levels below 5 are usually close to background noise levels.

Probe level QC: Log2 probe data of all arrays were inspected in DataDesk before and after quantile normalization. Inspection involved examining scatterplots of pairs of arrays for signal homogeneity (i.e., high correlation and linearity of the bivariate plots) and looking at all pairs of correlation coefficients. XY plots of probe expression and signal variance were also examined. Probe level array data sets were organized into reaction groups. Arrays with probe data that were not homogeneous when compared to other arrays were flagged.

Probe set level QC: The final normalized individual array data were evaluated for outliers. This involved counting the number of times that the probe set value for a particular array was beyond two standard deviations of the mean. This outlier analysis was carried out using the PDNN, RMA and MAS5 transforms and outliers across different levels of expression. Arrays that were associated with an average of more than 8% outlier probe sets across all transforms and at all expression levels were eliminated. In contrast, most other arrays generated fewer than 5% outliers.

Validation of strains and sex of each array data set: A subset of probes and probe sets with a Mendelian pattern of inheritance were used to construct a expression correlation matrix for all arrays and the ideal Mendelian expectation for each strain constructed from the genotypes. There should naturally be a very high correlation in the expression patterns of transcripts with Mendelian phenotypes within each strain, as well as with the genotype strain distribution pattern of markers for the strain.

Sex of the samples was validated using sex-specific probe sets such as Xist and Dby.



Notes:

This study includes the following datasets:

  • Hippocampus Consortium M430v2 CXB (Oct05) MAS5
  • Hippocampus Consortium M430v2 CXB (Oct05) RMA
  • Hippocampus Consortium M430v2 CXB (Oct05) PDNN
  • Hippocampus Consortium M430v2 CXB (Dec05) PDNN
  • Hippocampus Consortium M430v2 CXB (Dec05) RMA
  • Hippocampus Consortium M430v2 (Jun06) RMA
  • Hippocampus Consortium M430v2 (Jun06) MAS5
  • Hippocampus Consortium M430v2 (Jun06) PDNN 


Experiment Type:

Pooled RNA samples (usually one pool of male hippocampii and one pool of female hippocampii) were prepared using standard protocols. Samples were processed using a total of 206 Affymetrix GeneChip Mouse Expression 430 2.0 short oligomer arrays (MOE430 2.0 or M430v2; see GEO platform ID GPL1261), of which 201 passed quality control and error checking. This particular data set was processed using the PDNN protocol. To simplify comparisons among transforms, PDNN values of each array were adjusted to an average of 8 units and a standard deviation of 2 units.



Contributor:
  • David C. Airey, Ph.D.
    Grant Support: Vanderbilt Institute for Integratie Genomics
    Department of Pharmacology
    david.airey at vanderbilt.edu
  • Lu Lu, M.D.
    Grant Support: NIH U01AA13499, U24AA13513
  • Fred H. Gage, Ph.D.
    Grant Support: Lookout Foundation
  • Dan Goldowitz, Ph.D.
    Grant Support: NIAAA INIA AA013503
    University of Tennessee Health Science Center
    Dept. Anatomy and Neurobiology
    email: dgold@nb.utmem.edu
  • Shirlean Goodwin, Ph.D.
    Grant Support: NIAAA INIA U01AA013515
  • Gerd Kempermann, M.D.
    Grant Support: The Volkswagen Foundation Grant on Permissive and Persistent Factors in Neurogenesis in the Adult Central Nervous System
    Humboldt-Universitat Berlin
    Universitatsklinikum Charite
    email: gerd.kempermann at mdc-berlin.de
  • Kenneth F. Manly, Ph.D.
    Grant Support: NIH P20MH062009 and U01CA105417
  • Richard S. Nowakowski, Ph.D.
    Grant Support: R01 NS049445-01
  • Glenn D. Rosen, Ph.D.
    Grant Support: NIH P20
  • Leonard C. Schalkwyk, Ph.D.
    Grant Support: MRC Career Establishment Grant G0000170
    Social, Genetic and Developmental Psychiatry
    Institute of Psychiatry,Kings College London
    PO82, De Crespigny Park London SE5 8AF
    L.Schalkwyk@iop.kcl.ac.uk
  • Guus Smit, Ph.D.
    Dutch NeuroBsik Mouse Phenomics Consortium
    Center for Neurogenomics & Cognitive Research
    Vrije Universiteit Amsterdam, The Netherlands
    e-mail: guus.smit at falw.vu.nl
    Grant Support: BSIK 03053
  • Thomas Sutter, Ph.D.
    Grant Support: INIA U01 AA13515 and the W. Harry Feinstone Center for Genome Research
  • Stephen Whatley, Ph.D.
    Grant Support: XXXX
  • Robert W. Williams, Ph.D.
    Grant Support: NIH U01AA013499, P20MH062009, U01AA013499, U01AA013513


Citation:

Please cite: Overall RW, Kempermann G, Peirce J, Lu L, Goldowitz D, Gage FH, Goodwin S, Smit AB, Airey DC, Rosen GD, Schalkwyk LC, Sutter TR, Nowakowski RS, Whatley S, Williams RW (2009) Genetics of the hippocampal transcriptome in mice: a systematic survey and online neurogenomic resource. Front. Neurogen. 1:3 Full Text HTML doi:10.3389/neuro.15.003.2009



Data source acknowledgment:

Data were generated with funds provided by a variety of public and private source to members of the Consortium. All of us thank Muriel Davisson, Cathy Lutz, and colleagues at the Jackson Laboratory for making it possible for us to add all of the CXB strains, and one or more samples from KK/HIJ, WSB/Ei, NZO/HILtJ, LG/J, CAST/Ei, PWD/PhJ, and PWK/PhJ to this study. We thank Yan Cui at UTHSC for allowing us to use his Linux cluster to align all M430 2.0 probes and probe sets to the mouse genome. We thank Hui-Chen Hsu and John Mountz for providing us BXD tissue samples, as well as many strains of BXD stock. We thanks Douglas Matthews (UMem in Table 1) and John Boughter (JBo in Table 1) for sharing BXD stock with us. Members of the Hippocampus Consortium thank the following sources for financial support of this effort:

  • David C. Airey, Ph.D.
    Grant Support: Vanderbilt Institute for Integratie Genomics
    Department of Pharmacology
    david.airey at vanderbilt.edu
  • Lu Lu, M.D.
    Grant Support: NIH U01AA13499, U24AA13513
  • Fred H. Gage, Ph.D.
    Grant Support: Lookout Foundation
  • Dan Goldowitz, Ph.D.
    Grant Support: NIAAA INIA AA013503
    University of Tennessee Health Science Center
    Dept. Anatomy and Neurobiology
    email: dgold@nb.utmem.edu
  • Shirlean Goodwin, Ph.D.
    Grant Support: NIAAA INIA U01AA013515
  • Gerd Kempermann, M.D.
    Grant Support: The Volkswagen Foundation Grant on Permissive and Persistent Factors in Neurogenesis in the Adult Central Nervous System
    Humboldt-Universitat Berlin
    Universitatsklinikum Charite
    email: gerd.kempermann at mdc-berlin.de
  • Kenneth F. Manly, Ph.D.
    Grant Support: NIH P20MH062009 and U01CA105417
  • Richard S. Nowakowski, Ph.D.
    Grant Support: R01 NS049445-01
  • Glenn D. Rosen, Ph.D.
    Grant Support: NIH P20
  • Leonard C. Schalkwyk, Ph.D.
    Grant Support: MRC Career Establishment Grant G0000170
    Social, Genetic and Developmental Psychiatry
    Institute of Psychiatry,Kings College London
    PO82, De Crespigny Park London SE5 8AF
    L.Schalkwyk@iop.kcl.ac.uk
  • Guus Smit, Ph.D.
    Dutch NeuroBsik Mouse Phenomics Consortium
    Center for Neurogenomics & Cognitive Research
    Vrije Universiteit Amsterdam, The Netherlands
    e-mail: guus.smit at falw.vu.nl
    Grant Support: BSIK 03053
  • Thomas Sutter, Ph.D.
    Grant Support: INIA U01 AA13515 and the W. Harry Feinstone Center for Genome Research
  • Stephen Whatley, Ph.D.
    Grant Support: XXXX
  • Robert W. Williams, Ph.D.
    Grant Support: NIH U01AA013499, P20MH062009, U01AA013499, U01AA013513

 



Study Id:
23

CITG WWW service initiated January, 1994 as The Portable Dictionary of the Mouse Genome and June 15, 2001 as WebQTL. This site is currently operated by Rob Williams, Lei Yan, Pjotr Prins, Zachary Sloan, Arthur Centeno. Design and code by Lei Yan, Zach Sloan, Kenneth Manly, Jintao Wang, Danny Arends, Pjotr Prins, Sam Ockman, Xiaodong Zhou, Christian Fernandez, Ning Liu, Rudi Alberts, Elissa Chesler, Evan G. Williams, Alexander G. Williams, Robert W. Williams, and colleagues. Python Powered Registered with Nif
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