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Sample GSM886329 Query DataSets for GSM886329
Status Public on Mar 07, 2012
Title H37 Rv3132-34 KO 0 days vs Rv3132-34 KO (complemented) hypoxia 4 day rep 2
Sample type RNA
 
Channel 1
Source name H37Rv No treatment
Organism Mycobacterium tuberculosis
Characteristics treatment: control
strain name: H37Rv-Delta-dosR::kan plus DosR
Treatment protocol Cell pellets were suspended in 1 ml TRIzol reagent (GIBCO BRL) and transferred to 2-ml screw cap tubes containing 0.5 ml 0.1-mm diameter zirconia/silica beads (BioSpec Products).
Growth protocol Clinical isolate 1254, 7H9 medium (supplemented with BSA, NaCl, glucose, and glycerol), 250-ml vented tissue culture flasks, 90 rpm shaking, and a starting culture density of OD 0.15 were used, unless otherwise indicated. RNA samples isolated from OD 0.15 cultures of the M. tuberculosis strain being assayed on the same day were used for the reference sample in each experiment. Cells were collected with a 4-min centrifugation step and frozen on dry ice.
Extracted molecule total RNA
Extraction protocol Cell debris was separated by a 45-s centrifugation. The supernatant was transferred to 2-ml Heavy Phase Lock Gel I tubes (Eppendorf) containing 300 ul chloroform, inverted rapidly for 15 s, and incubated 2 min. Samples were centrifuged for 5 min, and the aqueous phase was added to 270 ul isopropanol, followed by addition of 270 ul of the following mixture: 0.8 M sodium citrate and 1.2 M NaCl. Samples were incubated for 10 min at 4 C and centrifuged for 15 min at 4 C. The RNA pellets were washed with 1 ml 75% ethanol, centrifuged 5 min, and air-dried. After suspension of the RNA pellets in 90 ul water, 10 ul DNase I 10x buffer, and 6 U DNase I (Ambion) were added, and the samples were incubated for 30 min. Final purification of RNA was by RNeasy column (QIAGEN).
Label Cy3
Label protocol Both a PCR gene product microarray and a 70-mer oligonucleotide-based microarray (tuberculosis oligonucleotide set; QIAGEN) were used. Labeled cDNA was prepared as follows: 2 ug total RNA and 4.4ug of random oligonucleotide hexamers were incubated for 2 min at 98 C, cooled on ice, combined with Stratascript RTase buffer, 0.5 mM dA,G,CTP, 0.02 mM dTTP, 1.5 nmol Cy3 or Cy5-dUTP (Amersham Biosciences), and 1.8 ul Stratascript RTase (Stratagene) in a total volume of 25 ul, and incubated for 10 min at 25 C and 90 min at 42 C. cDNA was purified by microcon-10 (Amicon) filtration.
 
Channel 2
Source name CO NRP Day 4
Organism Mycobacterium tuberculosis
Characteristics treatment: NRP (Wayne Growth)/4 days
strain name: H37Rv-Delta-dosR::kan plus DosR
Treatment protocol Cell pellets were suspended in 1 ml TRIzol reagent (GIBCO BRL) and transferred to 2-ml screw cap tubes containing 0.5 ml 0.1-mm diameter zirconia/silica beads (BioSpec Products).
Growth protocol Clinical isolate 1254, 7H9 medium (supplemented with BSA, NaCl, glucose, and glycerol), 250-ml vented tissue culture flasks, 90 rpm shaking, and a starting culture density of OD 0.15 were used, unless otherwise indicated. RNA samples isolated from OD 0.15 cultures of the M. tuberculosis strain being assayed on the same day were used for the reference sample in each experiment. Cells were collected with a 4-min centrifugation step and frozen on dry ice.
Extracted molecule total RNA
Extraction protocol Cell debris was separated by a 45-s centrifugation. The supernatant was transferred to 2-ml Heavy Phase Lock Gel I tubes (Eppendorf) containing 300 ul chloroform, inverted rapidly for 15 s, and incubated 2 min. Samples were centrifuged for 5 min, and the aqueous phase was added to 270 ul isopropanol, followed by addition of 270 ul of the following mixture: 0.8 M sodium citrate and 1.2 M NaCl. Samples were incubated for 10 min at 4 C and centrifuged for 15 min at 4 C. The RNA pellets were washed with 1 ml 75% ethanol, centrifuged 5 min, and air-dried. After suspension of the RNA pellets in 90 ul water, 10 ul DNase I 10x buffer, and 6 U DNase I (Ambion) were added, and the samples were incubated for 30 min. Final purification of RNA was by RNeasy column (QIAGEN).
Label Cy5
Label protocol Both a PCR gene product microarray and a 70-mer oligonucleotide-based microarray (tuberculosis oligonucleotide set; QIAGEN) were used. Labeled cDNA was prepared as follows: 2 ug total RNA and 4.4ug of random oligonucleotide hexamers were incubated for 2 min at 98 C, cooled on ice, combined with Stratascript RTase buffer, 0.5 mM dA,G,CTP, 0.02 mM dTTP, 1.5 nmol Cy3 or Cy5-dUTP (Amersham Biosciences), and 1.8 ul Stratascript RTase (Stratagene) in a total volume of 25 ul, and incubated for 10 min at 25 C and 90 min at 42 C. cDNA was purified by microcon-10 (Amicon) filtration.
 
 
Hybridization protocol 10 ul of hybridization solution (labeled cDNA, 5 ug tRNA, 3.8x SSC, 0.27% SDS) was sealed under a coverslip with rubber cement and hybridized overnight at 65 C for the DNA microarray. Oligonucleotide microarrays were first prehybridized for 1 h in 5x SSC, 1% BSA, and 0.1% SDS and washed with H2O and isopropanol. After the prehybridization, 10 ul of hybridization solution (labeled cDNA, 5 ug tRNA, 2x SSC, 25% formamide, and 0.1% SDS) was hybridized overnight at 54 C.
Scan protocol Microarrays were scanned using a GenePix 4000A (Axon Instruments, Inc.). The intensities of the two dyes at each spot were quantified using ScanAlyze (M. Eisen, Lawrence Berkeley National Lab, Berkeley, CA).
Description Simple annotation: Stress, Strain Comparison
Image: http://smd.stanford.edu/MicroArray/gifs/2007-06/75655.gif
Data processing VALUE is Log (base 2) of the ratio of the mean of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm)
 
Submission date Mar 05, 2012
Last update date Mar 07, 2012
Contact name SMD Staff
E-mail(s) array@genome.stanford.edu
Phone 650-498-6012
URL http://genome-www5.stanford.edu/
Organization name Stanford Microarray Database (SMD)
Department Stanford University, School of Medicine
Street address 300 Pasteur Drive
City Stanford
State/province CA
ZIP/Postal code 94305
Country USA
 
Platform ID GPL9901
Series (1)
GSE8839 Inhibition of Respiration by Nitric Oxide Induces a Mycobacterium tuberculosis Dormancy Program

Data table header descriptions
ID_REF ID_REF
CH1I_MEAN Uncorrected Cy3 mean pixel intensity.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel
CH2I_MEAN Uncorrected Cy3 mean pixel intensity.; Type: integer; Scale: linear_scale; Channel: Cy5 channel
CH1B_MEDIAN Median intensities of background pixels of Cy3.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel
CH2B_MEDIAN Median intensities of background pixels of Cy3.; Type: integer; Scale: linear_scale; Channel: Cy5 channel
CH1D_MEAN The mean feature pixel intensity with the median background subtracted (channel 1).; Type: integer; Scale: linear_scale; Channel: Cy3 Channel
CH2D_MEAN The mean feature pixel intensity with the median background subtracted (channel 2).; Type: integer; Scale: linear_scale; Channel: Cy5 channel
CH1B_MEAN Mean intensities of background pixels of Cy3.; Type: integer; Scale: linear_scale; Background
CH2B_MEAN Mean intensities of background pixels of Cy5.; Type: integer; Scale: linear_scale; Background
PERGTBCH1I_1SD The percentage of feature pixels with intensities more than one standard deviation above the background pixel intensity, at wavelength 532 nm.; Type: integer; Scale: linear_scale
PERGTBCH2I_1SD The percentage of feature pixels with intensities more than one standard deviation above the background pixel intensity, at wavelength 635 nm.; Type: integer; Scale: linear_scale
PIX_RAT2_MEDIAN Contains median of Ch2PI-CH2B/Ch1PI-CH1B where Ch1PI & Ch2PI represent single pixel intensities.; Type: float; Scale: linear_scale
TOT_SPIX Count of the number of pixels in the spot.; Type: integer; Scale: linear_scale
TOT_BPIX Number of background pixels.; Type: integer; Scale: linear_scale
REGR The regression ratio of every pixel in a 2-feature-diameter circle around the center of the feature.; Type: float; Scale: linear_scale
CORR The correlation between channel1 (Cy3) & Channel 2 (Cy5) pixels within the spot, and is a useful quality control parameter. Generally, high values imply better fit & good spot quality.; Type: float; Scale: linear_scale
TOP Box top: int(((centerX - radius) - Xoffset) / pixelSize).; Type: integer; Scale: linear_scale
BOT Box bottom: int(((centerX + radius) - Xoffset) / pixelSize).; Type: integer; Scale: linear_scale
LEFT Box left: int(((centerY - radius) - yoffset) / pixelSize).; Type: integer; Scale: linear_scale
RIGHT Box right: int(((centerY + radius) - yoffset) / pixelSize); Type: integer; Scale: linear_scale
FLAG User defined spot flag (default 0).; Type: integer; Scale: linear_scale
CH2IN_MEAN Normalized value of mean Channel 2 (usually 635 nm) intensity (CH2I_MEAN/Normalization factor).; Type: integer; Scale: linear_scale; Channel: Cy5 channel
CH2BN_MEDIAN Normalized value of median Channel 2 (usually 635 nm) background (CH2B_MEDIAN/Normalization factor).; Type: integer; Scale: linear_scale; Channel: Cy5 channel; Background
CH2DN_MEAN Normalized value of mean Channel 2 (usually 635 nm) intensity with normalized background subtracted (CH2IN_MEAN - CH2BN_MEDIAN).; Type: integer; Scale: linear_scale; Channel: Cy5 channel
RAT2N_MEAN Type: float; Scale: linear_scale
RAT1N_MEAN Ratio of the means of Channel 1 (usually 532 nm) intensity to normalized Channel 2 (usually 635 nm) intensity with median background subtracted (CH1D_MEAN/CH2DN_MEAN). Channel 1/Channel 2 ratio normalized or Green/Red ratio normalized.; Type: float; Scale: linear_scale
VALUE Log (base 2) of the ratio of the mean of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm) [log (base 2) (RAT2N_MEAN)].; Type: float; Scale: log_base_2

Data table
ID_REF CH1I_MEAN CH2I_MEAN CH1B_MEDIAN CH2B_MEDIAN CH1D_MEAN CH2D_MEAN CH1B_MEAN CH2B_MEAN PERGTBCH1I_1SD PERGTBCH2I_1SD PIX_RAT2_MEDIAN TOT_SPIX TOT_BPIX REGR CORR TOP BOT LEFT RIGHT FLAG CH2IN_MEAN CH2BN_MEDIAN CH2DN_MEAN RAT2N_MEAN RAT1N_MEAN VALUE
1 4464 6003 749 326 3715 5677 null null 89 92 1.506 225 1296 1.496 .968 51 68 55 72 0 4886 265 4620 1.244 .804 .315
2 2185 2219 758 332 1427 1887 null null 76 81 1.331 225 1205 1.294 .969 51 68 79 96 0 1806 270 1536 1.076 .929 .106
3 6401 8621 764 329 5637 8292 null null 88 92 1.498 225 1201 1.449 .98 51 68 101 118 0 7016 268 6749 1.197 .835 .26
4 2046 1726 763 328 1283 1398 null null 76 84 1.101 225 1234 1.076 .955 51 68 125 142 0 1405 267 1138 .887 1.127 -.173
5 7345 10025 761 336 6584 9689 null null 77 82 1.464 256 1203 1.442 .978 50 68 151 169 0 8159 273 7886 1.198 .835 .26
6 2573 2661 754 337 1819 2324 null null 77 82 1.242 256 1165 1.221 .963 50 68 175 193 0 2166 274 1891 1.04 .962 .056
7 1564 885 755 327 809 558 null null 77 83 .668 225 1204 .638 .882 50 67 198 215 0 721 266 454 .562 1.78 -.832
8 2608 2883 758 332 1850 2551 null null 80 88 1.397 225 1237 1.336 .969 50 67 222 239 0 2347 270 2077 1.122 .891 .167
9 1972 1924 765 351 1207 1573 null null 80 87 1.291 225 1207 1.243 .953 50 67 246 263 0 1566 286 1280 1.06 .943 .085
10 1342 1063 765 355 577 708 null null 69 76 1.223 225 1227 1.182 .9 50 67 268 285 0 865 289 576 .998 1.002 -.002
11 1409 817 755 347 654 470 null null 70 72 .709 225 1269 .701 .875 50 67 295 312 0 665 282 382 .585 1.711 -.775
12 1225 763 760 355 465 408 null null 68 73 .868 225 1238 .79 .85 50 67 319 336 0 621 289 332 .714 1.401 -.487
13 2504 1930 774 359 1730 1571 null null 79 84 .895 225 1201 .846 .95 50 67 343 360 0 1571 292 1279 .739 1.353 -.436
14 1167 913 771 361 396 552 null null 60 79 1.355 225 1201 1.179 .867 50 67 365 382 0 743 294 449 1.134 .882 .181
15 2716 4400 758 361 1958 4039 null null 83 92 2.052 225 1241 1.981 .972 50 67 389 406 0 3581 294 3287 1.679 .596 .748
16 3503 4845 745 353 2758 4492 null null 84 92 1.634 225 1254 1.569 .973 50 67 413 430 0 3943 287 3656 1.326 .754 .407
17 3481 1920 742 343 2739 1577 null null 86 87 .577 225 1318 .552 .951 50 67 438 455 0 1562 279 1283 .468 2.134 -1.094
18 2195 1412 737 309 1458 1103 null null 81 87 .754 225 1196 .738 .939 75 92 55 72 0 1149 251 898 .616 1.624 -.7
19 3537 3863 743 319 2794 3544 null null 88 92 1.28 225 1143 1.258 .98 75 92 77 94 0 3144 260 2884 1.032 .969 .046
20 1093 637 748 327 345 310 null null 56 65 .856 225 1148 .789 .81 75 92 103 120 0 519 266 253 .732 1.366 -.45

Total number of rows: 4608

Table truncated, full table size 539 Kbytes.




Supplementary data files not provided
Processed data included within Sample table

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