NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM542691 Query DataSets for GSM542691
Status Public on May 03, 2011
Title Haemophilus ducreyi 35000HPcpxA relative to 35000HP sample 4
Sample type RNA
 
Channel 1
Source name Haemophilus ducreyi 35000HP
Organism [Haemophilus] ducreyi
Characteristics strain: 35000HP
genotype/variation: wild type
Biomaterial provider Maria Labandeira-Rey
Treatment protocol Cells are grown under normal conditions
Growth protocol Haemophilus ducreyi strain was resucitated from frozen stock on Chocolate agar plates, and incubated ON at 33C in a humidified atmosphere containing 95% air and 5% CO2. Columbia broth supplemented with 2.5% FCS is inoculated from the plate and cultures are grown in a 33C gyratory water bath at 100 rpm for 8 hrs
Extracted molecule total RNA
Extraction protocol Total RNA from 2-4 ml of culture was extracted using the Ribopure Bacteria kit (Ambion) following manufacturer's instructions. After isolation, RNA was treated twice with DNaseI (NEB) for 1 hr at 37C, and purified using RNeasy system (Qiagen). Samples were checked for residual DNA by PCR and re-treated if necessary. Quality assessment was performed using the Agilent 2100 Bioanalyzer (Agilent Technologies)
Label Cy5
Label protocol Twenty micrograms of total RNA for cells grown in Columbia broth for 8 hr were used to generate cDNA using the Amino Allyl cDNA Labeling kit ragents (Ambion). After reverse transcription, residual RNA was removed by alkaline treatment, followed by neutralization. The cDNAs were purified using the Qiaquick gel extraction columns (Qiagen). After purification, Cy5 dye (GE Healthcare) was added following manufacturer's instructions. The labeled cDNAs were cleaned by using YM-30 Microcon centrifugal devices (Millipore) and labeling efficiency was determined using a Nanodrop spectrophotometer.
 
Channel 2
Source name Haemophilus ducreyi 35000HPcpxA
Organism [Haemophilus] ducreyi
Characteristics strain: 35000HP
genotype/variation: cpxA deletion mutant
Biomaterial provider Maria Labandeira-Rey
Treatment protocol Cells are grown under normal conditions
Growth protocol Haemophilus ducreyi strain was resucitated from frozen stock on Chocolate agar plates, and incubated ON at 33C in a humidified atmosphere containing 95% air and 5% CO2. Columbia broth supplemented with 2.5% FCS is inoculated from the plate and cultures are grown in a 33C gyratory water bath at 100 rpm for 8 hrs
Extracted molecule total RNA
Extraction protocol Total RNA from 2-4 ml of culture was extracted using the Ribopure Bacteria kit (Ambion) following manufacturer's instructions. After isolation, RNA was treated twice with DNaseI (NEB) for 1 hr at 37C, and purified using RNeasy system (Qiagen). Samples were checked for residual DNA by PCR and re-treated if necessary. Quality assessment was performed using the Agilent 2100 Bioanalyzer (Agilent Technologies)
Label Cy3
Label protocol Twenty micrograms of total RNA for cells grown in Columbia broth for 8 hr were used to generate cDNA using the Amino Allyl cDNA Labeling kit ragents (Ambion). After reverse transcription, residual RNA was removed by alkaline treatment, followed by neutralization. The cDNAs were purified using the Qiaquick gel extraction columns (Qiagen). After purification, Cy3 dye (GE Healthcare) was added following manufacturer's instructions. The labeled cDNAs were cleaned by using YM-30 Microcon centrifugal devices (Millipore) and labeling efficiency was determined using a Nanodrop spectrophotometer.
 
 
Hybridization protocol Equal amounts of labeled cDNA from 35000HP or 35000HPcpxA were thoroughly mixed and used to hybridize microarray slides in Amersham microarray hybridization buffer. Hybridization was carried out at 50C for 16 hr in the dark.
Scan protocol After hybridization, the slides were washe in saline-sodium phosphate EDTA buffer and scanned with GenePix 4100A scanner and analyzed with GenePix Pro 5.0 software (Axon instruments)
Description n/a
Data processing Data was subjected to two types of normalization, a ratio-based normalization so that the mean or median intensities are the same across the array, and a non-linear locally weighed scatterplot smoothing (LOWESS) normalization, which corrects intensity dependent variation in dye basis, before being combined into a single data set for further analysis. Differential expression was defined as a minimum of two fold over or under expression of 35000HPcpxA relative to 35000HP. The data were further scrutinized so as to only include expression profiles that were observed in at least 4 of the 6 experiments and had a p<0.05 after one t-test analysis.
 
Submission date May 11, 2010
Last update date May 03, 2011
Contact name Eric J Hansen
E-mail(s) eric.hansen@utsouthwestern.edu
Phone 2146331386
Organization name University of Texas Southwestern Medical Center
Department Microbiology
Lab Eric J. Hansen
Street address 5323 Harry Hines Blvd
City Dallas
State/province TX
ZIP/Postal code 75390-9048
Country USA
 
Platform ID GPL7741
Series (1)
GSE21788 Haemophilus ducreyi 35000HPcpxA gene expression relative to 35000HP

Data table header descriptions
ID_REF
VALUE Log2 ratio of the medians (35000HPcpxA/35000HP)
F532 Median 35000HPcpxA Signal median
B532 Median 35000HPcpxA Background median
F635 Median 35000HP Signal median
B635 Median 35000HP Background median

Data table
ID_REF VALUE F532 Median B532 Median F635 Median B635 Median
1:01:01 0.35831564 24.317 21.885 35.778 33.31
1:02:01 0.182414652 25.938 21.885 39.479 33.31
1:03:01 null 21.885 22.696 34.544 34.544
1:04:01 0.182414652 24.317 22.696 37.012 34.544
1:05:01 0.182414652 23.506 22.696 35.778 34.544
1:06:01 null 23.506 22.696 34.544 34.544
1:07:01 -0.118615343 23.506 22.696 37.012 34.544
1:08:01 -0.118615343 24.317 23.506 38.245 35.778
1:09:01 0.182414652 26.748 23.506 40.713 35.778
1:10:01 -0.118615343 26.748 23.506 45.648 35.778
1:11:01 0.182414652 25.938 23.506 39.479 35.778
1:12:01 -0.136082623 34.043 24.317 66.621 35.778
1:13:01 0.13893394 39.717 24.317 61.686 35.778
1:14:01 0.483444648 35.664 24.317 45.648 37.012
1:15:01 0.279438788 36.475 24.317 51.816 37.012
1:16:01 0.404320467 44.581 24.317 55.517 37.012
1:01:02 0.404320467 38.907 22.696 49.349 34.544
1:02:02 0.458939862 50.254 22.696 56.751 34.544
1:03:02 0.483444648 32.422 22.696 41.947 34.544
1:04:02 0.483444648 24.317 22.696 35.778 34.544

Total number of rows: 5760

Table truncated, full table size 262 Kbytes.




Supplementary file Size Download File type/resource
GSM542691.gpr.gz 701.9 Kb (ftp)(http) GPR
Processed data included within Sample table

| NLM | NIH | GEO Help | Disclaimer | Accessibility |
NCBI Home NCBI Search NCBI SiteMap