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Summary
Used the Affymetrix M430A and M430B pair of arrays (total of 45,137 probe sets). Data available as CEL files from GeneNetwork upon request.
About cases
The lines of mice used in this NCI-sponsored project consist of 18 groups of isogenic F1 progeny made by crossing females from each of 18 AKXD recombinant inbred strains (AKXD2, 3, 7, 9, 10, 11, 13, 14, 16, 18, 20, 21, 22, 23, 24, 25, 27, and 28) to male FVB/N mice that carry a transgene that consistently leads to the development of mammary tumors in females (e.g. Le Voyer et al., 2001). The formal nomenclature of the male transgenic line is FVB/N-TgN(MMTV-PyMT)634Mul. The genomes of each AKXD x FVB F1 consist of one set of FVB chromosomes (including the transgene) and one set of chromosomes inherited from one of the 18 AKXD RI strain mothers. Only the AKXD chromosomes are "recombinant" across this panel of F1 progeny, and the set therefore has a genetic architecture similar to backcross progeny. It is possible to map modifiers that influence tumor characteristics and expression patterns. It is also possible to study covariance of transcript expression levels in tumor tissue. For further background on this special mapping design please see Hunter and Williams (2002).
The ancestral strains from which all AKXD strains are derived are AKR/J (AKR) and DBA/2J (D2 or D). DBA/2J has been partially sequenced (approximately 1.5x coverage by D by Celera Genomics). Significant genomic sequence data for AKR is not currently available. Chromosomes of the two parental strains have recombined in the different AKXD strains. All of these strains are available from The Jackson Laboratory as cryopreserved stocks. For additional background on recombinant inbred strains, please see http://www.nervenet.org/papers/bxn.html.
About tissue
Mammary tumors used in this array experiment were derived from 18 sets of AKXD x FVB/N F1 females as described above. After the primary tumor was diagnosed, the animals were aged an additional 40 days to permit metastatic progression. Females were sacrificed and mammary tumors were harvested. Samples were processed and arrayed on Affymetrix M430A and M430B arrays. The majority of the samples were assayed on arrays obtained from the same lot number.
About platform
All samples were processed and arrayed in the Laboratory of Population Genetics at the NCI. The table below lists the arrays by Samples, AKXD strain and Age.
Sample
AKXD strain
Age
AKXD2957
2
98
AKXD2959
2
96
AKXD1747
3
84
AKXD3446
3
91
AKXD4225
3
83
AKXD2543
7
82
AKXD2967
7
88
AKXD3336
7
95
AKXD2685
9
113
AKXD2710
9
109
AKXD2949
9
115
AKXD2618
10
99
AKXD2620
10
99
AKXD3023
10
94
AKXD1910
11
87
AKXD2824
11
92
AKXD2825
11
103
AKXD2635
13
83
AKXD2718
13
100
AKXD2721
13
91
AKXD2632
14
99
AKXD2640
14
100
AKXD3444
14
96
AKXD1636
16
112
AKXD3688
16
80
AKXD4152
16
91
AKXD1647
18
91
AKXD2616
18
91
AKXD2804
18
80
AKXD2456
20
100
AKXD2554
20
107
AKXD2829
20
105
AKXD1610
21
98
AKXD2611
21
88
AKXD2918
21
98
AKXD2460
22
107
AKXD2461
22
94
AKXD2463
22
110
AKXD2975
23
82
AKXD2976
23
86
AKXD3955
23
90
AKXD1494
24
103
AKXD1880
24
104
AKXD3030
24
89
AKXD1607
25
110
AKXD2326
25
92
AKXD2328
25
90
AKXD2629
25
96
AKXD1756
27
100
AKXD1757
27
98
AKXD1948
27
99
AKXD1950
27
97
AKXD2968
27
94
AKXD2809
28
88
AKXD2815
28
90
AKXD3432
28
91
About data processing
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.Probe set data from the .CHP file: The expression data were generated using MAS5. The same simple steps described above were also applied to these values. A 1-unit difference represents roughly a two-fold difference in expression level. Expression levels below 5 are usually close to background noise levels.
- Step 1: We added an offset of 1.0 to the .CEL expression values for each cell to ensure that all values could be logged without generating negative values.
- Step 2: We took the log base 2 of each cell.
- Step 3: We computed the Z scores for each cell.
- Step 4: We multiplied all Z scores by 2.
- Step 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 corresponds approximately to a 1 unit difference.
- Step 6a: The 430A and 430B GeneChips include a set of 100 shared probe sets (2200 probes) that have identical sequences. These probes and probe sets provide a way to calibrate expression of the two GeneChips to a common scale. The absolute mean expression on the 430B array is almost invariably lower than that on the 430A array. To bring the two arrays into alignment, we regressed Z scores of the common set of probes to obtain a linear regression corrections to rescale the 430B arrays to the 430A array. In our case this involved multiplying all 430B Z scores by the slope of the regression and adding or subtracting a very small offset. The result of this step is that the mean of the 430A GeneChip expression is fixed at a value of 8, whereas that of the 430B chip is typically 7. Thus average of A and B arrays is approximately 7.5.
- Step 6b: We recenter the whole set of 430A and B transcripts to a mean of 8 and a standard deviation of 2. This involves reapplying Steps 3 through 5 above but now using the entire set of probes and probe sets from a merged 430A and B data set.
About the chromosome and megabase position values:
The chromosomal locations of probe sets and gene markers on the 430A and 430B microarrays were determined by BLAT analysis using the Mouse Genome Sequencing Consortium May 2004 (mm5) assembly (see http://genome.ucsc.edu/cgi-bin/hgBlat?command=start&org=mouse). We thank Dr. Yan Cui (UTHSC) for allowing us to use his Linux cluster to perform this analysis.
Acknowledgment
All of the NCI mammary mRNA M430A and M430B data sets have been generated by Kent Hunter at the Laboratory of Population Genetics at the National Cancer Institute in Bethesda. For contact and citations and other information on these data sets please review the INFO pages and contact Dr. Hunter regarding use of this data set in publications or projects.
Notes
Text originally written by Kent Hunter and Robert W. Williams, July 2004. Updated by RWW, Nov 6, 2004.