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Sample GSM2396041 Query DataSets for GSM2396041
Status Public on Nov 19, 2016
Title Mutant_type _ER79ap_with_ethanol_vs_no_ethanol (Replicate B)
Sample type RNA
 
Channel 1
Source name ethanol stress
Organism Zymomonas mobilis subsp. mobilis ZM4 = ATCC 31821
Characteristics strain: ZM4
genotype: mutant ER79ap
Growth protocol Zymomonas mobilis ZM4 and derivative ER79ap were grown in the medium reported by Bringer et al. (1985) but in wich the potassium phosphates were substituted by MES (100 mM, pH6). This modified medium was designated as MR-MES. Liquid cultures were performed in 350-ml minifermenters, containing 200 ml of MR-MES, without aeration, at 30° C, 100 rpm, and an initial pH of 6. The inoculum was prepared from a fresh colony obtained from solid MR-MES medium (agar 15 g/L). A single colony was seed in a test tube containing 10 ml of culture medium and incubated overnight (30° C and 250 rpm). Enough culture volume was used to start mini-fermenter cultures at 0.025 DO. Ethanol was added at 70 g/L.
Extracted molecule total RNA
Extraction protocol Total RNA was extracted from mid-log phase cells (with and without 70 g/L of ethanol) with a Quick-RNATM miniPrep (ZymoResearch, CA, USA) as described by the manufacturer
Label Alexa555
Label protocol 10 µg of total RNA were used for cDNA synthesis incorporanting dUTP-Alexa555 or DUTP-Alexa647 employing the Firs-Strand cDNA labeling kit (Invitrogen). Incorporation of fluorophore was analyzed by using the aobsorbance at 555 nm for Alexa555 and 650 nm for Alexa647. The sample ZM4_without Ethanol was labeled with Alexa555 and the sample ZM4_with Ethanol was labeled with Alexa647. The sample ER79ap_without Ethanol was labeled with Alexa555 and the sample ER79ap_with Ethanol was labeled with Alexa647
 
Channel 2
Source name none ethanol stress
Organism Zymomonas mobilis subsp. mobilis ZM4 = ATCC 31821
Characteristics strain: ZM4
genotype: mutant ER79ap
Growth protocol Zymomonas mobilis ZM4 and derivative ER79ap were grown in the medium reported by Bringer et al. (1985) but in wich the potassium phosphates were substituted by MES (100 mM, pH6). This modified medium was designated as MR-MES. Liquid cultures were performed in 350-ml minifermenters, containing 200 ml of MR-MES, without aeration, at 30° C, 100 rpm, and an initial pH of 6. The inoculum was prepared from a fresh colony obtained from solid MR-MES medium (agar 15 g/L). A single colony was seed in a test tube containing 10 ml of culture medium and incubated overnight (30° C and 250 rpm). Enough culture volume was used to start mini-fermenter cultures at 0.025 DO. Ethanol was added at 70 g/L.
Extracted molecule total RNA
Extraction protocol Total RNA was extracted from mid-log phase cells (with and without 70 g/L of ethanol) with a Quick-RNATM miniPrep (ZymoResearch, CA, USA) as described by the manufacturer
Label Alexa647
Label protocol 10 µg of total RNA were used for cDNA synthesis incorporanting dUTP-Alexa555 or DUTP-Alexa647 employing the Firs-Strand cDNA labeling kit (Invitrogen). Incorporation of fluorophore was analyzed by using the aobsorbance at 555 nm for Alexa555 and 650 nm for Alexa647. The sample ZM4_without Ethanol was labeled with Alexa555 and the sample ZM4_with Ethanol was labeled with Alexa647. The sample ER79ap_without Ethanol was labeled with Alexa555 and the sample ER79ap_with Ethanol was labeled with Alexa647
 
 
Hybridization protocol Zymomonas mobilis subsp. mobilis ZM4 has 1998 genes. MYcoarray have designed 5102 probes surveying 1894 genes (or 94.8% of all genes) for this organism. The array format was 3x15K, this slide contains 3 arrays of 15000 spots per array. One array consists of a grid of 160 columns by 94 rows. The siza of probe was 45mer. The Zymomonas mobilis chips from Mycroarray were re-hydrated with water vapor at 60° C, and fixed with two cycles of UV light (1200J). Equal quantities of labeled cDNA were hybridized using hybridization solution UniHyb (TeleChem International INC). The arrays were incubated for 14 h at 42° c, and then washed tree times with 1X SCC, 0.05% SDS at room temperature.
Scan protocol Acquisition and quantification of array images was performed in GenePix 4100A with its accompanying software GenePix form Molecular Devices. All images were captured using 10 µ resolution. For each spot the Alexa555 and Alexa647 density mean value and background mean value were calculated with software ArrayPro Analyzer from Media Cibernetics.
Description Biological replicate 2 of 3. (Mutant ER79ap ethanol vs ER79ap no ethanol stress)
http://berry.engin.umich.edu/oligoarraydb/organismPage.php?ORG=Zymomonas%20mobilis%20subsp.%20mobilis%20ZM8
Data processing Microarray data analysis was performed with free sofware genArise, developed in the Computing Unit of Cellular Physiology Institute of UNAM (http://www.ifc.unam.mx/genarise/). GenArise carry out a number of transformations: background correction, lowess normalization, intensity filter, replicates anlaysis and selecting differentially expressed genes. The goal of genArise is to identify wich of the gene show good evidence of being differentially expressed. The sofware identifies differential expressed genes by calculating an intensity-dependent z-score. Using a sliding window algorithm to calculate the mean and standard deviation within a window surrounding each data point, and define a z-score where z measures the number of strandard deviation a data point is form the mean. zi= (Ri - mean(R)) / sd (R) Where zi is the z-score for each element, Ri is the log-ratio for each element, and sd(R) is the standar deviation of the log-ratio. With this criterion, the elements with a z-score > 2 standard deviation would be the significantly differntially expressed genes.
With the software genArise, is to identify wich of the gene show good evidence of being differentially expressed. The sofware identifies differential expressed genes by calculating an intensity-dependent z-score. Using a sliding window algorithm to calculate the mean and standard deviation within a window surrounding each data point, and define a z-score where z measures the number of strandard deviation a data point is form the mean. zi= (Ri - mean(R)) / sd (R) Where zi is the z-score for each element, Ri is the log-ratio for each element, and sd(R) is the standar deviation of the log-ratio. With this criterion, the elements with a z-score > 1.5 standard deviation would be the significantly differntially expressed genes.
 
Submission date Nov 18, 2016
Last update date Nov 19, 2016
Contact name Rosa Maria Gutierrez Rios
E-mail(s) rmariab001@gmail.com
Phone (52) 777 3291634
Organization name UNIVERSIDAD NACIONAL AUTONOMA DE MEXICO
Department Molecular Microbiology
Lab Rosa Maria Gutierrez Rios
Street address AV. UNIVERSIDAD 3000, UNAM, CU
City MEXICO DISTRITO FEDERAL
State/province DISTRITO FEDERAL
ZIP/Postal code 04510
Country Mexico
 
Platform ID GPL22693
Series (1)
GSE90043 Phenotypic, genomic and transcriptional analysis of Zymomonas mobilis ZM4 mutants with enhanced ethanol tolerance

Data table header descriptions
ID_REF
VALUE Z-score

Data table
ID_REF VALUE
2774797 0.551077473
2774779 -0.385838746
2774815 0.210090313
2774838 0.193300609
2774832 1.058043713
2774826 0.916779477
2774782 0.955028556
2774800 0.382379738
2774918 0.605957099
2774854 0.691442161
2774829 -1.038384884
2774785 -1.429948491
2774794 -1.118737346
2774876 1.00894918
2774791 -0.246907312
2774788 0.633895195
2774806 -0.656184222
2774803 0.00543617
2774823 -0.280454848
2774809 -0.11354164

Total number of rows: 1886

Table truncated, full table size 37 Kbytes.




Supplementary file Size Download File type/resource
GSM2396041_raw_data_ER79apB.xls.gz 1.9 Mb (ftp)(http) XLS
Processed data included within Sample table

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