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Sample GSM590297 Query DataSets for GSM590297
Status Public on Dec 15, 2010
Title 55C_REP2
Sample type RNA
 
Source name human oocyte cDNA library 55°C
Organism Homo sapiens
Characteristics library: Human GV full-length oocyte cDNA library
hybridization temperature: 55C
Extracted molecule total RNA
Extraction protocol Total RNA was isolated from 14 GV using PicoPure RNA isolation kit (Molecular Devices, Sunnyvale, CA) following manufacturer's instruction. DNase treatment was performed with Rnase-free DNase I (Qiagen, Missisauga, On, Canada). RNA quantification and integrity were assessed with a 2100 BioAnalyzer apparatus (Agilent, Mississauga, On, Canada) using the RNA 6000 Picochip a Bioanalyzer (Agilent). The full-length human oocyte cDNA library was generated using the Creator SMART cDNA Library Construction Kit (Clontech, Mountain View, CA). a unique human probe was generated from plasmid library by PCR amplification using 50X Advantage 2 polymerase Mix (Clontech), and primers corresponding to pDNR-LIB sequence (Up : 5’- AGTCGACGGTACCGGACATA-3’, Low : 5’-GCCAAACGAATGGTCTAGAAA-3’) for 17 thermal cycles.
Label Alexa-647
Label protocol 2µg of cDNAs Purified PCR were then labelled using ULS Labeling Kit Alexafluor 547 and 647 dye pack (Kreatech, Amsterdam, The Netherlands); Diagnostics) according to manufacturer's instructions and unincorporated dyes were removed with Qiaquick PCR purification kit.
 
Hybridization protocol Sample probe was hybridized in technical replicate (2 replicates single-channel, on two slides) in SlideHyb™ Glass Array Hybridization Buffer #1 (Ambion, Streetville, Canada) for 18 hours at 45, 50, and 55°C respectively. Glass slides were then washed respectably at either 45, 50, or 55°C twice with 2X SCC/0.5% SDS for 15 min, twice with 0.5X SSC/0.5% SDS for 15 min, and rapidly washed in 1X SSC and H2O.
Scan protocol Slides were scanned using the VersArray ChipReader System (Bio-Rad) at 5 microns resolution and analyzed using the ChipReader and ArrayPro Analyzer software (Media Cybernetics, San Diego, CA).
Description Library was constructed with RNA extracted from 14 GV oocytes (CREATOR SMART, Clontech)
13724628_55°C_647
Data processing Data processed/normalized with ChipReader and ArrayPro Analyzer software (Media Cybernetics, San Diego, CA). Data were first normalized by Loess (by replicate within a same T°C), and signal was defined as present if intensity was above the threshold, calculated as negative and exogen DNA control cells mean intensity plus 1x standard deviation (GFP, plant and Alien DNA (Stratagene), SSC) for each array.
 
Submission date Sep 03, 2010
Last update date Dec 15, 2010
Contact name Eve-Lyne Sylvestre
E-mail(s) eve-lyne.sylvestre.1@ulaval.ca
Organization name Université Laval
Street address 2440 Boul Hochelaga
City Quebec
State/province Quebec
ZIP/Postal code G1V 0A6
Country Canada
 
Platform ID GPL10879
Series (1)
GSE23963 Hybridization of human oocyte cDNA library on the multi-species oocyte array to assess for gene conservation using a temperature stringency criteria

Data table header descriptions
ID_REF
VALUE ARRAY-PRO normalized (Loess) data signal intensity - (mean + 1SD intensity of negative and exogen control cells)

Data table
ID_REF VALUE
1 4376.621052
2
3 6085.126188
4 3331.861391
5 4149.329338
6 -143.007455
7 -105.08536
8
9 1126.308465
10 -47.516966
11 15321.02608
12 3720.532764
13 -114.377173
14
15 -153.83872
16
17 -153.375171
18 -146.113263
19 4359.133729
20

Total number of rows: 13824

Table truncated, full table size 221 Kbytes.




Supplementary data files not provided
Processed data included within Sample table

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