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Sample GSM14983 Query DataSets for GSM14983
Status Public on Jan 31, 2004
Title h15A
Sample type RNA
 
Channel 1
Source name D. melanogaster Canton S adult females
Organism Drosophila melanogaster
Extracted molecule total RNA
 
Channel 2
Source name adult hybrid females resulting from the cross D. melanogaster Canton S females x D. simulans Sim-1 males
Organism Drosophila melanogaster
Extracted molecule total RNA
 
 
Description Strains of Drosophila melanogaster and Drosophila simulans were raised on glucose-cornmeal-yeast medium at 25 C. Males and virgin females were collected for the interspecific and conspecific crosses. All crosses were performed at 18C using 7-8 individuals from each sex. Virgin females from the parental species as well as hybrid females were stored separately for 5-6 days at room temperature and then snap frozen in liquid nitrogen invariably at the same time of the day in a time window of 2 hours. For each cross, two independent cohorts were used as the source of mRNA. Total RNA was extracted by homogenizing 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 Jan 07, 2004
Last update date Jun 12, 2013
Contact name Jose M Ranz
E-mail(s) jmr68@mole.bio.cam.ac.uk
Phone 44 1223 766336
URL http://www.gen.cam.ac.uk/
Organization name University of Cambridge
Department Genetics
Lab Michael Ashburner
Street address Downing
City Cambridge
ZIP/Postal code cb2 3eh
Country United Kingdom
 
Platform ID GPL356
Series (1)
GSE948 whole body

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 three quality control criteria for inclusion of spots in analysis (1 = pass, 0 = failed), (1) [F635_MEAN-B635] > 2*[B635_SD] or [F532_MEAN-B532] > 2*[B532_SD], (2) spot passed PCR, (3) spot passed FLAGS
POLYMERASE polymerase used to amplify clone (0 - Taq [Amersham], 1 - Herculase [Stratagene])
NORM_F635 [(F635_MEAN-B635)*F_pixels]*SUM[(F532_MEAN-B532)*F_pixels]/SUM[(F635_MEAN-B635)*F_pixels] (SUM denotes the total intensity of fluorescence for a particular channel which is based only on those spots that passed QC_PASS; spots amplified by Taq and Herculase were normalized separately)
VALUE [NORM_F635]/[( F532_MEAN-B532)*F_pixels]

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
1.1.1 2695 12430 75 2153 2092 896 73 75 19 100 100 0 1374 1315 570 70 72 13 100 100 0 1.595 1.622 1.617 1.643 1.211 1.617 0.98 156 1272 3384 3264 0.674 2080 1304 2019 1245 0 1 1 0 138212.0116 0.711626051
1.1.2 2900 12430 75 1628 1495 709 72 75 20 100 100 0 422 394 151 70 71 12 100 99 0 4.42 4.392 4.477 4.26 1.384 4.552 0.944 156 1272 1908 1747 2.144 1556 352 1423 324 0 1 1 0 97412.42818 1.92727976
1.1.3 3100 12425 75 4900 4661 1134 73 76 29 100 100 0 1270 1206 282 69 71 11 100 100 0 4.019 4.035 4.042 4.062 1.176 4.062 0.978 156 1272 6028 5725 2.007 4827 1201 4588 1137 0 1 1 0 314074.6455 1.77071153
1.1.4 3295 12415 75 2608 2416 841 72 76 32 100 100 0 941 819 276 67 69 19 100 99 0 2.902 3.117 3.09 3.158 1.202 3.113 0.978 156 1246 3410 3096 1.537 2536 874 2344 752 0 1 1 0 160460.1066 1.367806419
1.1.5 3495 12420 85 13823 12061 5424 73 80 86 100 100 0 4351 3689 1526 67 69 18 100 100 0 3.21 3.31 3.333 3.213 1.277 3.378 0.985 208 1650 18034 15610 1.682 13750 4284 11988 3622 0 1 1 0 1094195.539 1.452389695
1.1.6 3695 12405 75 3141 2894 896 72 74 22 100 100 0 1263 1170 332 64 65 11 100 100 0 2.56 2.552 2.562 2.514 1.159 2.575 0.985 156 1202 4268 3928 1.356 3069 1199 2822 1106 0 1 1 0 193181.9201 1.119661521
1.1.7 3895 12410 75 25620 23527 7741 80 92 93 100 100 0 7914 7043 2400 70 79 80 100 100 0 3.256 3.363 3.339 3.437 1.141 3.34 0.987 156 1226 33384 30420 1.703 25540 7844 23447 6973 0 1 1 0 1605080.256 1.475545102
1.1.8 4090 12415 70 5424 4977 1966 78 80 21 100 100 0 1650 1503 549 67 74 92 100 98 0 3.377 3.412 3.465 3.388 1.165 3.462 0.986 156 1094 6929 6335 1.756 5346 1583 4899 1436 0 1 1 0 335364.361 1.497055393
1.1.9 4285 12420 70 2370 2380 928 76 77 19 100 100 0 1043 992 358 64 65 9 100 100 0 2.343 2.483 2.538 2.498 1.234 2.528 0.965 156 1114 3273 3232 1.228 2294 979 2304 928 0 1 1 0 157721.8795 1.089480268
1.1.10 4485 12420 80 6816 6570 4023 79 81 20 100 100 0 978 947 505 68 69 11 100 99 0 7.403 7.385 7.383 7.224 1.298 7.669 0.975 208 1484 7647 7370 2.888 6737 910 6491 879 0 0 0 0
1.1.11 4685 12410 75 8807 8582 1268 78 81 23 100 100 0 3482 3312 510 66 67 11 100 100 0 2.555 2.62 2.612 2.625 1.094 2.62 0.988 156 1202 12145 11750 1.354 8729 3416 8504 3246 0 1 1 0 582147.0761 1.149634019
1.1.12 4880 12425 80 7186 6474 3621 79 81 23 100 100 0 4439 3894 2147 67 69 14 100 100 0 1.626 1.671 1.667 1.701 1.34 1.687 0.973 208 1484 11479 10222 0.701 7107 4372 6395 3827 0 1 1 0 583698.7381 0.733275133
1.1.13 5085 12425 90 11186 10422 8156 84 92 46 99 95 0 4363 4013 3012 70 83 53 95 90 0 2.586 2.622 2.626 2.67 1.559 2.689 0.973 256 1894 15395 14281 1.371 11102 4293 10338 3943 0 0 0 0
1.1.14 5285 12420 80 5043 4089 2523 77 85 52 100 100 0 3130 2541 1518 67 73 44 99 98 0 1.621 1.622 1.64 1.597 1.345 1.643 0.984 208 1462 8029 6486 0.697 4966 3063 4012 2474 0 1 1 0 366192.2341 0.711616648
1.1.15 5480 12430 80 2395 2062 1093 80 86 44 98 97 0 3798 3277 1849 69 77 61 99 98 0 0.621 0.618 0.631 0.642 1.29 0.607 0.977 208 1502 6044 5190 -0.688 2315 3729 1982 3208 0 1 1 0 180905.5354 0.271115384
1.1.16 5675 12435 90 10089 10274 6694 93 118 78 100 100 0 1092 932 518 77 124 109 92 81 0 9.848 11.908 12.245 11.417 1.4 13.171 0.905 256 1924 11011 11036 3.3 9996 1015 10181 855 0 1 1 0 1143708.301 5.2252755
1.1.17 5880 12430 80 4482 3851 2397 104 123 72 98 95 0 857 762 395 86 129 99 85 80 0 5.678 5.543 5.503 4.897 1.374 5.906 0.957 208 1484 5149 4423 2.505 4378 771 3747 676 0 1 1 0 342004.5616 2.43232648
1.1.18 6080 12430 75 3733 3631 641 92 95 26 100 100 0 1399 1321 252 76 78 12 100 100 0 2.752 2.843 2.843 2.871 1.165 2.84 0.972 156 1262 4964 4784 1.461 3641 1323 3539 1245 0 1 1 0 242264.6404 1.247372261
1.1.19 6280 12435 75 5938 5135 2432 94 97 31 100 100 0 2083 1830 764 75 77 13 100 100 0 2.91 2.872 2.837 2.721 1.302 2.972 0.972 156 1272 7852 6796 1.541 5844 2008 5041 1755 0 1 1 0 345085.0671 1.260446589
1.1.20 6480 12430 70 8986 8268 2819 93 99 41 100 100 0 11021 9552 3458 74 83 63 100 99 0 0.812 0.863 0.857 0.899 1.288 0.848 0.978 156 1114 19840 17653 -0.3 8893 10947 8175 9478 0 1 1 0 559625.1584 0.378491323

Total number of rows: 6101

Table truncated, full table size 1181 Kbytes.




Supplementary data files not provided
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