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Sample GSM923055 Query DataSets for GSM923055
Status Public on Apr 26, 2012
Title Co-cultured K. vulgare Cells, Replicate 3
Sample type RNA
 
Channel 1
Source name Co-cultured K. vulgare cells
Organisms Priestia megaterium; Ketogulonicigenium vulgare WSH-001
Characteristics growth condition: co-culture with Bacillus megaterium
growth phase: mid-logarithmic
Treatment protocol The mixture of K. vulgare and B. megaterium was grown at 30 °C in the well-defined chemical medium at pH 7.0, 200 rpm for 18 h until cells were at their mid-log phase.
Growth protocol The mono-culture of K. vulgare was grown at 30 °C in the well-defined chemical medium at pH 7.0, 200rpm for 18 h.
Extracted molecule total RNA
Extraction protocol The mono- and co-cultures were sampled after 18-h fermentation (mid-log phase) and added to RNAProtect® Bacterial Reagent (Qiagen, Valencia, CA) according to the manufacturer’s instructions. Bacterial cells were treated with lysozyme stock solution (40 mg ml-1 in 30 mM pH 8.0 Tris/EDTA buffer) and protease K at 25oC for 10 min (vortex 10 s for every 2 min). Total RNA was isolated and purified using the RNeasy Mini kit (Qiagen) according to the manufacturer’s protocol. Total RNA binding on the RNeasy spin column membrane were treated with RNase-Free DNase Set (Qiagen) to remove remaining DNA. The concentration and purity of the total RNA were assessed by Nanodrop® ND-8000 (Thermo Fisher), and the integrity of the RNA was assessed using standard denaturing agarose gel electrophoresis.
Label Cy5
Label protocol Sample labeling and hybridization reactions were performed according to the Agilent Two-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technology). Briefly, for each sample, 1 μg of total RNA was linearly amplified and labeled with Cy5-dCTP and sample of common reference with Cy3-dCTP. Labelled cRNAs were purified with the Qiagen RNeasy Mini Kit, and sample concentration and specific activity (pmol Cy5/Cy3 per μg cRNA) were measured in a NanoDrop® ND-8000 spectrophotometer.
 
Channel 2
Source name Pooled 3 co-cultured and 3 mono-cultured cells
Organisms Priestia megaterium; Ketogulonicigenium vulgare WSH-001
Characteristics growth condition: co-culture with Bacillus megaterium and mono-culture
growth phase: mid-logarithmic
sample type: reference
Treatment protocol The mixture of K. vulgare and B. megaterium was grown at 30 °C in the well-defined chemical medium at pH 7.0, 200 rpm for 18 h until cells were at their mid-log phase.
Growth protocol The mono-culture of K. vulgare was grown at 30 °C in the well-defined chemical medium at pH 7.0, 200rpm for 18 h.
Extracted molecule total RNA
Extraction protocol The mono- and co-cultures were sampled after 18-h fermentation (mid-log phase) and added to RNAProtect® Bacterial Reagent (Qiagen, Valencia, CA) according to the manufacturer’s instructions. Bacterial cells were treated with lysozyme stock solution (40 mg ml-1 in 30 mM pH 8.0 Tris/EDTA buffer) and protease K at 25oC for 10 min (vortex 10 s for every 2 min). Total RNA was isolated and purified using the RNeasy Mini kit (Qiagen) according to the manufacturer’s protocol. Total RNA binding on the RNeasy spin column membrane were treated with RNase-Free DNase Set (Qiagen) to remove remaining DNA. The concentration and purity of the total RNA were assessed by Nanodrop® ND-8000 (Thermo Fisher), and the integrity of the RNA was assessed using standard denaturing agarose gel electrophoresis.
Label Cy3
Label protocol Sample labeling and hybridization reactions were performed according to the Agilent Two-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technology). Briefly, for each sample, 1 μg of total RNA was linearly amplified and labeled with Cy5-dCTP and sample of common reference with Cy3-dCTP. Labelled cRNAs were purified with the Qiagen RNeasy Mini Kit, and sample concentration and specific activity (pmol Cy5/Cy3 per μg cRNA) were measured in a NanoDrop® ND-8000 spectrophotometer.
 
 
Hybridization protocol A total of 1μg of Cy3-labeled and Cy5-labeled cRNA was prepared for fragmentation adding 11 μl 10 × Blocking Agent and 2.2 μl of 25× Fragmentation Buffer, heated at 60 °C for 30 min, and finally diluted by addition of 55μl 2× GE Hybridization buffer. A volume of 100 μl of hybridization solution was then dispensed in the gasket slide and assembled to the gene expression microarray slide. The slides were incubated for 17 h at 65°C in an Agilent Hybridization Oven.
Scan protocol The hybridized arrays were washed, fixed and scanned using the Agilent DNA microarray scanner (part number G2505B) following Agilent Technology’s guidelines.
Description Co-cultured-rep3
Common reference design.
Data processing Agilent Feature Extraction Software (version 10.5.1.1) was used to analyze acquired array images. Lowess normalization and subsequent data processing was performed using the Agilent GeneSpring GX v11.5.1 software. Following Lowess normalization of the raw data, genes for which at least 8 out of 16 samples have flags in Present or Marginal (“All Targets Value”) were chosen for further data analysis. Differentially expressed genes with statistical significance were identified through Volcano Plot filtering. The threshold we used to screen up- or downregulated genes was defined by Fold Change ≥2 and P-value <0.05.
 
Submission date Apr 25, 2012
Last update date Apr 26, 2012
Contact name Zhu Yibo
E-mail(s) centuryrain@126.com
Organization name Jiangnan University
Street address Lihu Road 1800#
City Wuxi
State/province Jiangsu
ZIP/Postal code 214122
Country China
 
Platform ID GPL14693
Series (1)
GSE37594 Ketogulonigenium vulgare cells: co-culture mode vs. mono-culture mode

Data table header descriptions
ID_REF
VALUE Lowess-normalized log2 ratio (test/reference)

Data table
ID_REF VALUE
KVU_0001 -0.3473835
KVU_0002 -1.7829399
KVU_0003 -0.7692766
KVU_0004 -0.5481186
KVU_0009 -0.08261299
KVU_0012 -0.8117943
KVU_0013 -0.21814251
KVU_0014 -0.5086813
KVU_0024 -0.26392508
KVU_0029 0.11421585
KVU_0031 -0.13753223
KVU_0034 0.17660236
KVU_0038 -1.5794306
KVU_0039 -0.7870827
KVU_0040 0.1236372
KVU_0041 0.11579037
KVU_0043 0.054876328
KVU_0044 -0.27508354
KVU_0050 -0.11746979
KVU_0051 0.18789482

Total number of rows: 1234

Table truncated, full table size 24 Kbytes.




Supplementary file Size Download File type/resource
GSM923055_CKH3.txt.gz 1.4 Mb (ftp)(http) TXT
Processed data included within Sample table

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