NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM495675 Query DataSets for GSM495675
Status Public on Jun 13, 2012
Title H37Rv dosR KO + pMVdosR vs dosR KO rep 8
Sample type RNA
 
Channel 1
Source name H37Rv MRO dosR KO
Organism Mycobacterium tuberculosis
Characteristics strain: H37Rv
genotype: MRO dosR KO
Extracted molecule total RNA
Extraction protocol TB-Synthesis and Labeling of cDNA SKlab 01; 1. Bring 0.5-5 ?g RNA (Typically 3 ?g) to 11 ?l and add 2 ?l (~2 mg/ml) random hexamers. 2. Heat 10 min at 65C (or 2 min at 98C), snap cool on ice. 10 rxns 20 rxns 3. Add 11?l 5.0 ?l 5X First-Strand buffer (51) (102) 2.5 ?l 100 mM DTT (25.5) (51) 2.0 ?l dNTP (5 mM A,G,C and 0.2 mM T) (20.5) (41) 1.5 ?l Cy3 or Cy5 (Typically use Cy5 for sample that should change) (15.3) (30.6) Then add 1.2-1.8 ?l Stratascript or 0.8-1.2 ?l Superscript II RTase. 4. Incubate 10m at 25C followed by 90m at 42C in PCR machine. Can freeze or leave at 4C o/n.; Protocol Type = Labeling; Performer: Mario Alberto,,Flores-Valdez
Label Cy3
Label protocol not provided
 
Channel 2
Source name H37Rv MRO (dosRKO) + pMVdosR
Organism Mycobacterium tuberculosis
Characteristics strain: H37Rv
experiment: MRO (dosRKO) + pMVdosR
Growth protocol Growth Conditions; Parameters that describe the conditions under which the biological sample was grown or propagated; Protocol Type = Growth Conditions; Parameter OD at Harvest = 0.2; Parameter Growth Temperature = 37oC; Parameter Media = Middlebrook 7H9 supplemented with albumin-dextrose complex (ADC); Performer: Mario Alberto,,Flores-Valdez
Extracted molecule total RNA
Extraction protocol TB-RNA Isolation and Purification SKlab 01; 1. Centrifuge 20-30 ml of early- to mid-log culture (O.D. 0.1-0.2) 4 min at 3700 rpm, at RT-37C. Pipette off supernatant and immediately freeze on dry ice and store at ?80C. 2. Add 1 ml Trizol (Gibco-BRL) to each of 4-8 pellets, suspend first pellet by vortexing,while other pellets are still frozen. Add suspension to 0.4 ml glass beads in a 2 ml screw cap tube. 3. Shake 30s at maximum speed in bead beater. Suspend next pellet and add to glass beads. Apply next sample to bead beater. Repeat bead beating two more times with each sample.Continue to periodically invert samples for at least 5 min in Trizol. 4. Centrifuge samples 45s at max speed, remove Trizol solution to a 2 ml screw cap tube containing half of the Heavy Phase Lock Gel I, supplied by 5 Prime 3 Prime, Inc. in 1.5 ml tubes and transferred with sterile stick to 2 ml tubes, and 300 ?l Chloroform:isoamyl alcohol (24:1). Invert rapidly for 15s, and continue inverting periodically for 2 min.a 5. Centrifuge 5-10m, remove aqueous layer (540 ?l) and add to a 1.5 ml tube containing 270 ?l isopropanol then add 270 ?l high salt solution (0.8M Na Citrate, 1.2M NaCl). Invert several times and spray with Staphene and remove from the P3. Typically ppted at 4C O/N. 6. Centrifuge 10m at 4C and remove isopropanol. Add 1 ml 75% EtOHb, invert several times and centrifuge 5m. 7. Remove EtOH by aspiration. Then, dry under vacuum for 2 min [Do not over dry]. 8. Suspend RNA in 90 ml RNase free water (don?t suspend in DNaseI buffer), may need to heat 10m at 55-60C to dissolve RNA. (Optional: obtain RNA concentration) 9. Add 10 ?l 10X DNaseI buffer to RNA (use no more than 80 ?g RNA) and add 4 ?l DNaseI (Ambion). Incubate 30m at 37C. Qiagen RNeasy purification. 1. Add 350 ?l RLT buffer (add 10 ?l BME to 1 ml RLT before using) and vortex. Add 260 ?l 95% EtOH (or 250 ?l 100% EtOH) to each sample and vortex. 2. Add to RNeasy spin column, centrifuge 15s, transfer column to a new 2 ml collection tube. Add 500 ?l RPE, centrifuge 15s, discard flow-through, add 500 ?l additional RPE and centrifuge 2m. If column still wet on sides, remove wash solution from tube and spin 1m to dry. 3. Transfer to a 1.5 ml collection tube, elute with 40 ?l RNase free water, centrifuge 1m. 4. Determine RNA concentration with A260/A280 readings. Dilute 1 ?l in 199 ?l TE. [200(dilution factor)x 40 ?g/A260x A260 = ?g/ml]. 5. Run 1 ?l RNA on 2% Agarose TAE gel. Run gel 45m at 100 volts. aIf Phase Lock Gel is not used, decrease Chloroform:isoamyl alcohol to 200?l. The yield will be less. Be careful not to remove any of the interface layer. bUsing 100% EtOH should be avoided in all steps of preparation of RNA intended for array analysis. Instead use 95% EtOH in preparation of solutions. Benzene contamination may fluoresce. cInstead of using the RNeasy purification, the DNaseI can be inactivated at 65-70C for 15m and then EtOH precipitated. The RNeasy purification may not be necessary in all applications. We find the purification gives more consistent results and is less time consuming, compared to precipitation. In addition, other purifications can be used. Note: The RNeasy column will remove much of the small tRNA and degraded RNA.; Protocol Type = Extract preparation; Performer: Mario Alberto,,Flores-Valdez
Label Cy5
Label protocol Amount of extract labeled = 2-3 ug of total RNA
 
 
Hybridization protocol not provided
Scan protocol GenePix
Description Simple annotation: Absolute transcript levels, Profiling
Image: http://smd.stanford.edu/MicroArray/gifs/2005-10/64958.gif
Data processing VALUE is Log (base 2) of the ratio of the median of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm)
 
Submission date Jan 11, 2010
Last update date Jun 13, 2012
Contact name Mario A Flores Valdez
E-mail(s) floresvz@yahoo.com
Organization name Centro de Investigación y Asistencia en Tecnología y diseño del Estado de Jalisco, A.C.
Department Col. Colinas de la Normal
Street address Av. Normalistas No. 800
City Guadalajara
State/province Jalisco
ZIP/Postal code CP44270
Country Mexico
 
Platform ID GPL9901
Series (1)
GSE19838 Gene expression profiles of M. tuberculosis constitutively expressing the transcriptional regulator DosR (Rv3133c).

Data table header descriptions
ID_REF ID_REF
CH1I_MEAN Mean feature pixel intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale
CH2I_MEAN Mean feature pixel intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale
CH1B_MEDIAN The median feature background intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel; Background
CH2B_MEDIAN The median feature background intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Channel: Cy5 channel; Background
CH1D_MEAN The mean feature pixel intensity at wavelength 532 nm with the median background subtracted.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel
CH2D_MEAN .The mean feature pixel intensity at wavelength 635 nm with the median background subtracted.; Type: integer; Scale: linear_scale; Channel: Cy5 channel
CH1I_MEDIAN Median feature pixel intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale
CH2I_MEDIAN Median feature pixel intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale
CH1B_MEAN The mean feature background intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale; Background
CH2B_MEAN The mean feature background intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Background
CH1D_MEDIAN The median feature pixel intensity at wavelength 532 nm with the median background subtracted.; Type: integer; Scale: linear_scale
CH2D_MEDIAN The median feature pixel intensity at wavelength 635 nm with the median background subtracted.; Type: integer; Scale: linear_scale
CH1_PER_SAT The percentage of feature pixels at wavelength 532 nm that are saturated.; Type: integer; Scale: linear_scale
CH2_PER_SAT The percentage of feature pixels at wavelength 635 nm that are saturated.; Type: integer; Scale: linear_scale
CH1I_SD The standard deviation of the feature intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel
CH2I_SD The standard deviation of the feature pixel intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Channel: Cy5 channel
CH1B_SD The standard deviation of the feature background intensity at wavelength 532 nm.; Type: float; Scale: linear_scale; Channel: Cy3 Channel; Background
CH2B_SD The standard deviation of the feature background intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Channel: Cy5 channel; 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
PERGTBCH1I_2SD The percentage of feature pixels with intensities more than two standard deviations above the background pixel intensity, at wavelength 532 nm.; Type: integer; Scale: linear_scale
PERGTBCH2I_2SD The percentage of feature pixels with intensities more than two standard deviations above the background pixel intensity, at wavelength 532 nm.; Type: integer; Scale: linear_scale
SUM_MEAN The sum of the arithmetic mean intensities for each wavelength, with the median background subtracted.; Type: integer; Scale: linear_scale
SUM_MEDIAN The sum of the median intensities for each wavelength, with the median background subtracted.; Type: integer; Scale: linear_scale
RAT1_MEAN Ratio of the arithmetic mean intensities of each spot for each wavelength, with the median background subtracted. Channel 1/Channel 2 ratio, (CH1I_MEAN - CH1B_MEDIAN)/(CH2I_MEAN - CH2B_MEDIAN) or Green/Red ratio.; Type: float; Scale: linear_scale
RAT2_MEAN The ratio of the arithmetic mean intensities of each feature for each wavelength, with the median background subtracted.; Type: float; Scale: linear_scale
RAT2_MEDIAN The ratio of the median intensities of each feature for each wavelength, with the median background subtracted.; Type: float; Scale: linear_scale
PIX_RAT2_MEAN The geometric mean of the pixel-by-pixel ratios of pixel intensities, with the median background subtracted.; Type: float; Scale: linear_scale
PIX_RAT2_MEDIAN The median of pixel-by-pixel ratios of pixel intensities, with the median background subtracted.; Type: float; Scale: linear_scale
RAT2_SD The geometric standard deviation of the pixel intensity ratios.; Type: float; Scale: linear_scale
TOT_SPIX The total number of feature pixels.; Type: integer; Scale: linear_scale
TOT_BPIX The total 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
DIAMETER The diameter in um of the feature-indicator.; Type: integer; Scale: linear_scale
X_COORD X-coordinate of the center of the spot-indicator associated with the spot, where (0,0) is the top left of the image.; Type: integer; Scale: linear_scale
Y_COORD Y-coordinate of the center of the spot-indicator associated with the spot, where (0,0) is the top left of the image.; Type: integer; 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 The type of flag associated with a feature: -100 = user-flagged null spot; -50 = software-flagged null spot; 0 = spot valid.; 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
CH2IN_MEDIAN Normalized value of median Channel 2 (usually 635 nm) intensity (CH2I_MEDIAN/Normalization factor).; Type: integer; Scale: linear_scale
CH2DN_MEDIAN Normalized value of median Channel 2 (usually 635 nm) intensity with normalized background subtracted (CH2IN_MEDIAN - CH2BN_MEDIAN).; Type: integer; 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
RAT2N_MEDIAN Channel 2/Channel 1 ratio normalized, RAT2_MEDIAN/Normalization factor or Red/Green median ratio normalized.; Type: float; Scale: linear_scale
LOG_RAT2N_MEAN 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
VALUE Log (base 2) of the ratio of the median of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm) [log (base 2) (RAT2N_MEDIAN)].; Type: float; Scale: log_base_2

Data table
ID_REF CH1I_MEAN CH2I_MEAN CH1B_MEDIAN CH2B_MEDIAN CH1D_MEAN CH2D_MEAN CH1I_MEDIAN CH2I_MEDIAN CH1B_MEAN CH2B_MEAN CH1D_MEDIAN CH2D_MEDIAN CH1_PER_SAT CH2_PER_SAT CH1I_SD CH2I_SD CH1B_SD CH2B_SD PERGTBCH1I_1SD PERGTBCH2I_1SD PERGTBCH1I_2SD PERGTBCH2I_2SD SUM_MEAN SUM_MEDIAN RAT1_MEAN RAT2_MEAN RAT2_MEDIAN PIX_RAT2_MEAN PIX_RAT2_MEDIAN RAT2_SD TOT_SPIX TOT_BPIX REGR CORR DIAMETER X_COORD Y_COORD TOP BOT LEFT RIGHT FLAG CH2IN_MEAN CH2BN_MEDIAN CH2DN_MEAN RAT2N_MEAN CH2IN_MEDIAN CH2DN_MEDIAN RAT1N_MEAN RAT2N_MEDIAN LOG_RAT2N_MEAN VALUE
1 428 295 37 37 391 258 439 301 38 38 402 264 0 0 102 81 7 8 100 100 100 98 649 666 1.516 .66 .657 .643 .659 1.539 156 992 .654 .95 140 1410 28280 46 60 30 44 0 483 61 423 1.081 493 432 .925 1.075 .112 .105
2 133 72 37 38 96 34 134 74 38 38 97 36 0 0 25 17 6 8 100 91 100 87 130 133 2.824 .354 .371 .321 .362 1.935 156 922 .365 .82 140 1650 28270 45 59 54 68 0 118 62 56 .58 121 59 1.724 .608 -.786 -.718
3 313 145 37 38 276 107 327 147 38 39 290 109 0 0 78 36 6 8 100 99 100 98 383 399 2.579 .388 .376 .383 .389 1.642 156 952 .374 .92 140 1890 28270 45 59 78 92 0 237 62 175 .635 241 179 1.575 .616 -.655 -.7
4 142 74 37 38 105 36 144 72 38 38 107 34 0 0 23 16 6 8 100 94 100 88 141 141 2.917 .343 .318 .297 .336 2.081 156 921 .343 .8 140 2130 28270 45 59 102 116 0 121 62 59 .561 118 56 1.781 .52 -.833 -.942
5 677 383 37 37 640 346 674 373 38 38 637 336 0 0 145 85 6 8 100 100 100 100 986 973 1.85 .541 .527 .541 .535 1.352 156 938 .523 .94 140 2360 28270 45 59 125 139 0 627 61 567 .885 611 550 1.129 .864 -.176 -.211
6 251 129 36 37 215 92 259 129 37 37 223 92 0 0 46 28 5 7 100 100 100 100 307 315 2.337 .428 .413 .423 .423 1.381 156 937 .428 .92 140 2610 28270 45 59 150 164 0 211 61 151 .701 211 151 1.427 .676 -.513 -.566
7 117 64 37 36 80 28 119 63 37 37 82 27 0 0 23 15 5 8 99 91 99 78 108 109 2.857 .35 .329 .312 .323 1.847 156 937 .343 .75 140 2850 28270 45 59 174 188 0 105 59 46 .573 103 44 1.745 .539 -.803 -.891
8 219 125 37 37 182 88 216 125 37 37 179 88 0 0 27 24 5 8 100 100 100 100 270 267 2.068 .484 .492 .469 .495 1.328 120 838 .472 .92 130 3090 28260 44 57 198 211 0 205 61 144 .792 205 144 1.263 .805 -.337 -.313
9 124 64 37 36 87 28 125 64 37 37 88 28 0 0 22 15 6 7 99 92 98 79 115 116 3.107 .322 .318 .285 .327 2.171 156 954 .296 .72 140 3330 28270 45 59 222 236 0 105 59 46 .527 105 46 1.897 .521 -.924 -.94
10 104 60 36 36 68 24 103 58 37 37 67 22 0 0 17 13 5 8 100 88 100 66 92 89 2.833 .353 .328 .31 .323 1.854 156 1038 .382 .72 150 3570 28260 43 58 245 260 0 98 59 39 .578 95 36 1.73 .538 -.791 -.895
11 124 69 36 36 88 33 128 68 36 37 92 32 0 0 21 15 5 7 100 96 100 90 121 124 2.667 .375 .348 .342 .36 1.766 156 922 .372 .82 140 3810 28270 45 59 270 284 0 113 59 54 .614 111 52 1.628 .57 -.703 -.812
12 109 72 36 36 73 36 112 72 36 37 76 36 0 0 21 17 5 7 98 96 98 89 109 112 2.028 .493 .474 .469 .467 2.099 156 951 .474 .77 140 4040 28270 45 59 293 307 0 118 59 59 .808 118 59 1.238 .776 -.308 -.366
13 194 94 35 37 159 57 194 92 37 37 159 55 0 0 25 18 15 7 100 100 100 100 216 214 2.789 .358 .346 .345 .35 1.444 120 850 .334 .85 130 4290 28260 44 57 318 331 0 154 61 93 .587 151 90 1.703 .566 -.768 -.82
14 98 54 36 36 62 18 98 53 36 37 62 17 0 0 16 13 5 7 100 78 99 58 80 79 3.444 .29 .274 .263 .279 2.125 156 954 .303 .67 140 4530 28270 45 59 342 356 0 88 59 29 .475 87 28 2.103 .449 -1.073 -1.155
15 242 153 36 36 206 117 244 153 37 37 208 117 0 0 36 29 9 8 100 100 100 100 323 325 1.761 .568 .563 .558 .578 1.362 120 812 .559 .91 130 4760 28270 45 58 365 378 0 251 59 192 .93 251 192 1.075 .921 -.104 -.118
16 392 251 37 37 355 214 408 259 38 38 371 222 0 0 85 64 8 8 100 99 100 98 569 593 1.659 .603 .598 .579 .612 1.79 156 969 .604 .94 140 5000 28270 45 59 389 403 0 411 61 350 .987 424 364 1.013 .98 -.019 -.029
17 308 202 36 36 272 166 332 201 36 37 296 165 0 0 73 56 5 7 100 100 100 100 438 461 1.639 .61 .557 .61 .617 1.515 156 1020 .608 .94 140 5250 28270 45 59 414 428 0 331 59 272 .999 329 270 1.001 .913 -.001 -.131
18 166 110 38 39 128 71 165 113 39 40 127 74 0 0 23 25 7 8 100 100 100 99 199 201 1.803 .555 .583 .523 .564 1.46 120 797 .563 .9 130 1420 28500 68 81 31 44 0 180 64 116 .908 185 121 1.101 .954 -.139 -.068
19 284 141 37 38 247 103 282 137 38 39 245 99 0 0 46 34 7 8 100 99 100 98 350 344 2.398 .417 .404 .394 .416 1.47 156 885 .423 .92 140 1660 28500 68 82 55 69 0 231 62 169 .683 224 162 1.464 .662 -.55 -.596
20 90 55 38 37 52 18 87 54 38 38 49 17 0 0 16 15 6 8 100 70 100 51 70 66 2.889 .346 .347 .302 .339 2.425 120 774 .336 .55 130 1900 28510 69 82 79 92 0 90 61 29 .567 88 28 1.764 .568 -.819 -.816

Total number of rows: 4608

Table truncated, full table size 931 Kbytes.




Supplementary data files not provided
Processed data included within Sample table
| NLM | NIH | GEO Help | Disclaimer | Accessibility |
NCBI Home NCBI Search NCBI SiteMap