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Sample GSM7884 Query DataSets for GSM7884
Status Public on Jul 16, 2003
Title E87a
Sample type RNA
 
Channel 1
Source name Zimbabwe30 adult males
Organism Drosophila melanogaster
Extracted molecule total RNA
 
Channel 2
Source name Canton S adult males
Organism Drosophila melanogaster
Extracted molecule total RNA
 
 
Description Strains of Drosophila melanogaster were raised on standard medium at 25 C with no light/dark cycle. Adult males were collected up to 24 hours after eclosing and separated from females where they were allowed to age an additional 3-4 days. Total RNA was extracted by homogenizing 8-12 flies in TRIzol reagent (Invitrogen) followed by a chloroform extraction and isopropanol precipitation. The resulting pellet was washed with 75% ethanol, air-dried, and resuspended in TE. Individual extractions were pooled into one common RNA sample for each strain. Poly-A RNA was purified from total RNA using the Oligotex Direct mRNA kit (Qiagen). mRNA samples were confirmed to be of high quality with a 2100 Bioanalyzer (Agilent). The production of cDNA and hybridizations were done following a published protocol (Eisen and Brown, Methods in Enzymology 303:179-205). 2 µg of poly-A RNA was used as a template for SuperScript II reverse transcriptase (Invitrogen) in the presence of amino-allyl dUTP (Sigma) with random hexamer and oligo dT primers. Reverse transcription was done at 42 C for 3 hours, followed by degradation of RNA by the addition of NaOH and EDTA and incubation at 65 C for 15 minutes. The cDNA was concentrated and purified with Microcon-30 filters (Millipore). Cyanine-3 or cyanine-5 fluorochromes (Amersham) were incorporated with the addition of 1 M NaHCO3 and a 75 minute incubation at room temperature with the cDNA probe. The labeled probes were then purified using the QIAquick PCR Purification Kit (Qiagen), concentrated with Microcon-30 filters, and combined into one sample. 25 µL of probe was combined with 22 µg poly(A), 4.4 µL 20X SSC, 0.71 µL 1 M HEPES (pH 7.0), and 0.66 µL 10% SDS. The probe was boiled at 100 C for 2 minutes and injected under a LifterSlip (Erie Scientific Company) placed over the array. Slides were placed in a Telechem hybridization chamber with 3X SSC in the chamber wells to prevent drying, sealed, and placed in a 65 C water bath for 12 hours. After hybridization, slides were washed with two wash solutions (0.6X SSC, 0.025% SDS [solution 1] and 0.05X SSC [solution 2]). Slides were dried by centrifugation at 1000 rpm for 2 minutes and scanned within an hour of washing. Fluorescence intensity was measured with Axon GenePix 4000A or 4000B scanners and the data was captured with GenePix 4.0 software. Following exclusion of suboptimal spots, intensity ratios were constructed by subtracting local background from foreground for each spot and normalizing one channel so that the mean foreground fluorescence of both channels across the slide was equal.
 
Submission date Jul 14, 2003
Last update date Mar 20, 2006
Contact name Colin Meiklejohn
E-mail(s) [email protected]
Phone 617-496-5540
Fax 617-496-5854
URL http://www.oeb.harvard.edu/hartl/lab/
Organization name Harvard University
Department Organismic and Evolutionary Biology
Lab Hartl Lab
Street address 16 Divinity Ave
City Cambridge
State/province MA
ZIP/Postal code 02138
Country USA
 
Platform ID GPL356
Series (1)
GSE539 Intraspecific variation in gene expression

Data table header descriptions
ID_REF
X x coordinate position in scanned area of slide (center of spot)
Y y coordinate position in scanned area of slide (center of spot)
DIA diameter of spot
F635_MED Cy5 foreground median pixel intensity
F635_MEAN Cy5 foreground mean pixel intensity
F635_SD Cy5 foreground pixel intensity standard deviation
B635_MED Cy5 background median pixel intensity
B635_MEAN Cy5 background mean pixel intensity
B635_SD Cy5 background pixel intensity standard deviation
%>B635+1SD percent of foreground pixels with a Cy5 intensity greater than (background + 1 standard deviation of background)
%>B635+2SD percent of foreground pixels with a Cy5 intensity greater than (background + 2 standard deviations of background)
F635%SAT percentage of foreground pixels saturated in the Cy5 channel
F532_MED Cy3 foreground median pixel intensity
F532_MEAN Cy3 foreground mean pixel intensity
F532_SD Cy3 foreground pixel intensity standard deviation
B532_MED Cy3 background median pixel intensity
B532_MEAN Cy3 background mean pixel intensity
B532_SD Cy3 background pixel intensity standard deviation
%>B532+1SD percent of foreground pixels with a Cy3 intensity greater than (background + 1 standard deviation of background)
%>B532+2SD percent of foreground pixels with a Cy3 intensity greater than (background + 2 standard deviations of background)
F532%SAT percentage of foreground pixels saturated in the Cy3 channel
RATIO_OF_MEDIANS(635/532) ratio of median pixel intensities, with each pixel intensity corrected by subtracting the median background intensity for that channel
RATIO_OF_MEANS(635/532) ratio of mean pixel intensities, with each pixel intensity corrected by subtracting the median background intensity for that channel
MEDIAN_OF_RATIOS(635/532) median ratio of pixel intensities
MEAN_OF_RATIOS(635/532) mean ratio of pixel intensities
RATIOS_SD(635/532) standard deviation of ratios of pixel intensities
RGN_RATIO(635/532) slope of the least-squares fit linear regression between the two channels, including foreground and background pixels
RGN_R^2(635/532) R^2 for the linear regression
F_PIXELS number of foreground pixels in spot
B_PIXELS number of background pixels in spot
SUM_OF_MEDIANS sum of the background corrected median pixel intensity for Cy5 and Cy3
SUM_OF_MEANS sum of the background corrected mean pixel intensity for Cy5 and Cy3
LOG_RATIO(635/532) log2 of the ratio of medians
F635_MED-B635 Cy5 foreground median intensity minus Cy5 background median intensity
F532_MED-B532 Cy3 foreground median intensity minus Cy3 background median intensity
F635_MEAN-B635 Cy5 foreground mean intensity minus Cy5 background median intensity
F532_MEAN-B532 Cy3 foreground mean intensity minus Cy3 background median intensity
FLAGS quality control identifiers (100 or 0 = good spot, -50 = spot not found by imaging software, -100 = spot flagged as bad by user)
PCR agarose gel quality control of PCR amplification of cDNAs (1 = pass, 0 = fail)
QC_PASS six quality control criteria for inclusion of spots in analysis (1 = pass, 0 = failed), (1) [F635_MEAN-B635] > 3*[B635_SD] or [F532_MEAN-B532] > 3*[B532_SD], (2) [%>B635+2SD] > 70 and [%>B532+2SD] > 70, (3) [F635%SAT] < 10 and [F532%SAT] < 10, (4) [B635_MED] < 3*[B532_MED] and [B532_MED] < 3*[B635_MED], (5) spot passed PCR, (6) spot passed FLAGS
POLYMERASE polymerase used to amplify clone (0 - Taq [Amersham], 1 - Herculase [Stratagene])
NORM_F635 [F635_MEAN-B635]*mean[F532_MEAN-B532]/mean[F635_MEAN-B635] (means were calculated only including spots that passed QC_PASS, spots amplified by Taq and Herculase were normalized separately)
VALUE log ratio of PRE_VALUE
PRE_VALUE [NORM_F635]/[ F635_MEAN-B635]

Data table
ID_REF X Y DIA F635_MED F635_MEAN F635_SD B635_MED B635_MEAN B635_SD %>B635+1SD %>B635+2SD F635%SAT F532_MED F532_MEAN F532_SD B532_MED B532_MEAN B532_SD %>B532+1SD %>B532+2SD F532%SAT RATIO_OF_MEDIANS(635/532) RATIO_OF_MEANS(635/532) MEDIAN_OF_RATIOS(635/532) MEAN_OF_RATIOS(635/532) RATIOS_SD(635/532) RGN_RATIO(635/532) RGN_R^2(635/532) F_PIXELS B_PIXELS SUM_OF_MEDIANS SUM_OF_MEANS LOG_RATIO(635/532) F635_MED-B635 F532_MED-B532 F635_MEAN-B635 F532_MEAN-B532 FLAGS PCR QC_PASS POLYMERASE NORM_F635 VALUE PRE_VALUE
1.1.1 2855 12060 65 2010 1939 573 64 66 17 100 100 0 2803 2689 812 66 68 14 100 100 0 0.711 0.715 0.718 0.717 1.192 0.711 0.979 120 914 4683 4498 -0.492 1946 2737 1875 2623 0 1 1 0 1830.03861 -0.1563383 0.697689138
1.1.10 4655 12065 75 1303 1248 608 70 73 20 100 100 0 1246 1201 503 72 73 13 100 100 0 1.05 1.043 1.039 0.981 1.308 1.098 0.957 156 1262 2407 2307 0.071 1233 1174 1178 1129 0 0 0 0
1.1.11 4850 12065 85 6080 5402 3269 69 72 24 100 100 0 6467 5637 3205 71 75 29 100 100 0 0.94 0.958 0.962 0.915 1.26 0.978 0.983 208 1641 12407 10899 -0.09 6011 6396 5333 5566 0 1 1 0 5205.117818 -0.0291133 0.935163101
1.1.12 5050 12070 75 17800 15656 5782 71 75 29 100 100 0 18166 16276 5620 73 80 50 100 100 0 0.98 0.962 0.971 0.951 1.201 0.971 0.982 156 1262 35822 31788 -0.029 17729 18093 15585 16203 0 1 1 0 15211.28093 -0.0274303 0.938794108
1.1.13 5245 12070 75 11050 9438 4426 72 76 30 100 100 0 10272 9126 3885 75 79 31 100 100 0 1.077 1.035 1.048 0.992 1.227 1.072 0.977 156 1262 21175 18417 0.106 10978 10197 9366 9051 0 0 0 0
1.1.14 5450 12070 85 5247 4809 3141 73 77 25 100 100 0 4073 3917 2388 75 77 22 100 100 0 1.294 1.233 1.242 1.152 1.289 1.264 0.983 208 1650 9172 8578 0.372 5174 3998 4736 3842 0 1 1 0 4622.433524 0.0803133 1.203132099
1.1.15 5655 12070 70 4643 4668 1476 74 76 20 100 100 0 6102 6157 1811 75 77 16 100 100 0 0.758 0.755 0.763 0.749 1.157 0.76 0.983 156 1114 10596 10676 -0.4 4569 6027 4594 6082 0 1 1 0 4483.8386 -0.1323963 0.737230944
1.1.16 5850 12070 80 1169 1073 613 73 76 20 99 99 0 1267 1155 620 75 77 18 100 99 0 0.919 0.926 0.911 0.877 1.329 0.945 0.971 208 1484 2288 2080 -0.121 1096 1192 1000 1080 0 1 1 0 976.0205921 -0.0439653 0.90372277
1.1.17 6050 12065 75 1069 940 448 71 74 19 100 100 0 912 777 336 73 75 12 100 100 0 1.19 1.234 1.201 1.182 1.373 1.27 0.956 156 1246 1837 1573 0.25 998 839 869 704 0 1 1 0 848.1618945 0.0809063 1.204775418
1.1.18 6245 12075 90 1121 1365 1060 71 73 18 99 98 0 1048 1240 909 72 73 10 99 99 0 1.076 1.108 1.089 1.03 1.334 1.136 0.982 256 1912 2026 2462 0.105 1050 976 1294 1168 0 1 1 0 1262.970646 0.0339503 1.081310485
1.1.19 6440 12075 90 861 1677 1562 72 75 19 99 96 0 994 1663 1453 73 74 10 98 98 0 0.857 1.009 0.977 0.899 1.485 1.045 0.977 256 1912 1710 3195 -0.223 789 921 1605 1590 0 1 1 0 1566.51305 -0.0064633 0.985228334
1.1.2 3065 12070 65 840 817 201 65 67 17 100 100 0 812 791 187 66 67 10 100 100 0 1.039 1.037 1.036 1.038 1.193 1.046 0.956 120 940 1521 1477 0.055 775 746 752 725 0 1 1 0 733.9674853 0.0053393 1.012368945
1.1.20 6645 12070 85 9961 9386 7098 73 79 37 100 100 0 10876 10092 7142 76 80 41 100 100 0 0.916 0.93 0.916 0.844 1.343 0.956 0.981 208 1650 20688 19329 -0.127 9888 10800 9313 10016 0 1 1 0 9089.679774 -0.0421463 0.907515952
1.1.21 6845 12075 80 690 667 406 72 75 20 99 96 0 870 785 459 74 76 15 99 98 0 0.776 0.837 0.841 0.797 1.462 0.853 0.958 208 1492 1414 1306 -0.365 618 796 595 711 0 1 1 0 580.7322523 -0.0878943 0.816782352
1.1.3 3265 12060 65 2084 1957 424 65 68 18 100 100 0 2254 2222 388 68 70 13 100 100 0 0.924 0.878 0.879 0.867 1.168 0.898 0.967 120 914 4205 4046 -0.115 2019 2186 1892 2154 0 1 1 0 1846.63096 -0.0668663 0.857303138
1.1.4 3460 12070 75 2266 1827 812 65 69 23 100 100 0 2104 1760 734 68 69 11 100 100 0 1.081 1.041 1.029 1.003 1.248 1.056 0.971 156 1272 4237 3454 0.112 2201 2036 1762 1692 0 1 1 0 1719.748283 0.0070653 1.016399695
1.1.5 3665 12065 80 6149 5186 3272 67 69 17 100 100 0 7038 6331 3664 68 69 12 100 100 0 0.873 0.817 0.825 0.743 1.384 0.841 0.981 208 1502 13052 11382 -0.197 6082 6970 5119 6263 0 1 1 0 4996.249411 -0.0981383 0.797740605
1.1.6 3860 12065 85 1147 979 669 67 69 18 99 99 0 941 921 590 69 70 11 100 100 0 1.239 1.07 1.047 0.979 1.449 1.097 0.97 208 1650 1952 1764 0.309 1080 872 912 852 0 1 1 0 890.13078 0.0190143 1.044754437
1.1.7 4055 12070 80 9023 7724 5076 69 72 18 100 100 0 6964 6408 4027 70 72 15 100 100 0 1.299 1.208 1.211 1.158 1.39 1.231 0.977 208 1512 15848 13993 0.377 8954 6894 7655 6338 0 1 1 0 7471.437633 0.0714523 1.178832066
1.1.8 4255 12070 75 3686 3308 1640 69 73 22 100 100 0 3219 2842 1409 72 74 18 100 100 0 1.149 1.169 1.169 1.17 1.261 1.165 0.977 156 1272 6764 6009 0.201 3617 3147 3239 2770 0 1 1 0 3161.330698 0.0573903 1.14127462

Total number of rows: 6101

Table truncated, full table size 1240 Kbytes.




Supplementary data files not provided
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