|
| Status |
Public on Jan 10, 2006 |
| Title |
2h post-infection of THP-1 treated with 2 rounds of SCOTS, replicate 3 |
| Sample type |
mixed |
| |
|
| Channel 1 |
| Source Name |
2XSCOTS cDNA obtained 2h post-infection
|
| Organism(s) |
Salmonella typhi |
| Characteristics |
Salmonella typhi infection of THP-1 cells, recovered 2 h post-infection
|
| Treatment protocol |
2E7 THP-1 cells (treated with 1E-7 M phorbol 12-myristate 13-acetate) were infected with Salmonella typhi at a multiplicity of infection of 10:1. At time 0, cells were washed with PBS and fresh RPMI containing 100 ug ml-1 was added to the wells. After 2 hours, cells were washed with PBS and recovered in 5 ml PBS. After centrifuation, the pellet was lysed with 2 ml TRIzol (Invitrogen).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
TRIzol reagent (Gibco BRL) was used according to the manufacturer instruction. RNA was treated with DNase (Ambion). RNA sample was converted to cDNA in 5 independent reverse-transcription reactions as described previously (Graham, J. 1999. PNAS 96:11554-559). Bacterial transcript were purified from host transcript by using 2 round of the SCOTS procedure (Daigle, F. 2002. Methods Enzymol. 358:108-122).
|
| Label |
Cy5
|
| Label protocol |
1.5 µg of cDNA was labeled with Cy5-dCTP (GE Healthcare) by random priming with Klenow (New England Biolabs) as previously described (Porwollik, S. 2001. Mutat Res 483:1-11.).
|
| |
|
| Channel 2 |
| Source Name |
Reference genomic DNA
|
| Organism(s) |
Salmonella typhi |
| Characteristics |
Genomic DNA from Salmonella typhi ISP1820
|
| Growth protocol |
Salmonella typhi from overnight culture in LB broth.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was extracted by standard phenol-chloroform method (Sambrook, J, Molecular Cloning, 3rd Edition)
|
| Label |
Cy3
|
| Label protocol |
1.5 µg of genomic DNA was labeled with Cy3-dCTP (GE Healthcare) by random priming with Klenow (New England Biolabs) as previously described (Porwollik, S. 2001. Mutat Res 483:1-11.).
|
| |
|
| |
| Hybridization protocol |
Samples we hybridized as previously described (Porwolik, S. 2003.Nucleic Acids Res 31:1869-76)
|
| Scan protocol |
Slide was scanned with Scan Array Lite (Perkin-Elmer) using Scan Array Express 2.1 software.
|
| Description |
This sample was used to analyse the effect of SCOTS on transcript population.
|
| Data processing |
After backgroud correction and total intensity normalization, ratio to reference were calculated and log base 2 transformed.
|
| |
|
| Submission date |
Aug 09, 2005 |
| Contact name |
Sébastien P Faucher |
| E-mail(s) |
sebastien.faucher@umontreal.ca
|
| Phone |
514-343-6111 #3135
|
| Fax |
514-343-5701
|
| Organization name |
University of Montreal
|
| Department |
Microbiology and Immunology
|
| Street address |
C.P. 6128 Succ. Centre-Ville
|
| City |
Montreal |
| State/province |
QUEBEC |
| ZIP/Postal code |
H3C 3J7 |
| Country |
Canada |
| |
|
| Platform ID |
GPL548 |
| Series (1) |
| GSE3095 |
Analysis of SCOTS effect on transcript population |
|
| Data table header descriptions |
| ID_REF |
Number |
| X_LOCATION |
X-coordinate of spot |
| Y_LOCATION |
Y-coordinate of spot |
| CH1_MEAN |
Channel 1 mean |
| CH1_BKD_MEAN |
Channel 1 mean background |
| CH1_MEAN_SD |
|
| CH1_BKD_SD |
Channel 1 background standard deviation |
| CH1_DIAMETER |
Channel 1 diameter |
| CH1_AREA |
Channel 1 area |
| CH1_FOOTPRINT |
Channel 1 footprint |
| CH1_CIRCULARITY |
Channel 1 circularity |
| CH1_SPOT_UNIFORMITY |
Channel 1 spot uniformity |
| CH1_BKD_UNIFORMITY |
Channel 1 background uniformity |
| CH1_SIGNAL_NOISE_RATIO |
Channel 1 signal to noise ratio |
| CH1_CONFIDENCE |
Channel 1 confidence |
| CH2_MEAN |
Channel 2 mean |
| CH2_BKD_MEAN |
Channel 2 mean background |
| CH2_SD |
Channel 2 standard deviation |
| CH2_BKD_SD |
Channel 2 background standard deviation |
| CH2_DIAMETER |
Channel 2 diameter |
| CH2_AREA |
Channel 2 area |
| CH2_FOOTPRINT |
Channel 2 footprint |
| C2_CIRCULARITY |
|
| CH2_SPOT_UNIFORMITY |
Channel 2 spot uniformity |
| CH2_BKD_UNIFORMITY |
Channel 2 background uniformity |
| CH2_SIGNAL_NOISE_RATIO |
Channel 2 signal to noise ratio |
| CH2_CONFIDENCE |
Channel 2 confidence |
| CH1_DIAMETER_FILTER |
Channel 1 diameter filter |
| CH1_AREA_FILTER |
Channel 1 area filter |
| CH1_CIRCULARITY_FILTER |
Channel 1 circularity filter |
| CH1_SPOT_UNIFORMITY_FILTER |
Channel 1 spot uniformity filter |
| CH1_BKD_UNIFORMITY_FILTER |
Channel 1 background uniformity filter |
| CH1_SIGNAL_NOISE_RATIO_FILTER |
Channel 1 signal to noise ratio filter |
| CH2_DIAMETER_FILTER |
Channel 2 diameter filter |
| CH2_AREA_FILTER |
Channel 2 area filter |
| CH2_CIRCULARITY_FILTER |
Channel 2 circularity filter |
| CH2_SPOT_UNIFORMITY_FILTER |
Channel 2 spot uniformity filter |
| CH2_BKD_UNIFORMITY_FILTER |
Channel 2 background uniformity filter |
| CH2_SIGNAL_NOISE_RATIO_FILTER |
Channel 2 signal to noise ratio filter |
| IGNORE_FILTER |
Ignore filter |
| CH1_BKD_CORR |
Channel 1 background correlation |
| CH2_BKD_CORR |
Channel 2 background correlation |
| CH1%_OF_TOTAL |
Channel 1 % of total |
| CH2%_OF_TOTAL |
Channel 2 % of total |
| PRE-VALUE |
ch1 % of total / ch2 % of total |
| VALUE |
log base 2 of ch1 % of total / ch2 % of total |
