|
| Status |
Public on Jul 22, 2015 |
| Title |
cDNA array L20/H20 (dye swap) |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
20 DPA cotton fiber of Line 2
|
| Organism |
Gossypium hirsutum |
| Characteristics |
strain: Extracted total RNA from 20 DPA Line 2 cotton fiber
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using CTAB method
|
| Label |
Cy3
|
| Label protocol |
Double-stranded cDNAs were synthesized from 5 µg total RNA using the cDNA synthesis system following the manufacturer’s protocol (CapitalBio, China). A T7-oligo (dT) primer was used in this reaction.The cDNAs were mixed with 4 µl of random primer, heated to 95°C for 3 min and snap-cooled on ice for 5 min. 5 µl of 5 × klenow buffer, 1 µl Cy5-dCTP or Cy3-dCTP (GE Healthcare), and 1.2 µl of klenow fragment were added, and the reaction was kept at 37°C for 1.5 h, 70°C for 5 min and cooled on ice for 5 min. Labeled cDNA was purified using a PCR Nucleospin® Extraction kit (MN).
|
| |
|
| Channel 2 |
| Source name |
20 DPA cotton fiber of Line 1
|
| Organism |
Gossypium hirsutum |
| Characteristics |
strain: Extracted total RNA from 20 DPA Line 1 cotton fiber
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using CTAB method
|
| Label |
Cy5
|
| Label protocol |
Double-stranded cDNAs were synthesized from 5 µg total RNA using the cDNA synthesis system following the manufacturer’s protocol (CapitalBio, China). A T7-oligo (dT) primer was used in this reaction.The cDNAs were mixed with 4 µl of random primer, heated to 95°C for 3 min and snap-cooled on ice for 5 min. 5 µl of 5 × klenow buffer, 1 µl Cy5-dCTP or Cy3-dCTP (GE Healthcare), and 1.2 µl of klenow fragment were added, and the reaction was kept at 37°C for 1.5 h, 70°C for 5 min and cooled on ice for 5 min. Labeled cDNA was purified using a PCR Nucleospin® Extraction kit (MN).
|
| |
|
| |
| Hybridization protocol |
For microarray hybridization, labeled controls and test samples were quantitatively adjusted, based on efficiency of Cy-dye incorporation, and were mixed with 80 µl of hybridization solution (3 × SSC, 0.2% SDS, 5 × Denhart’s, 25% formamide). Arrays were hybridized at 42°C overnight and washed in two consecutive solutions (0.2% SDS, 2 × SSC at 42°C for 5 min, and 0.2 SSC for 5 min at room temperature).
|
| Scan protocol |
Scanned on a LuxScanTM 10 K microarray scanner (CapitalBio Corp.).
Images were quantified using LuxScan 3.0 software (CapitalBio Corp.).
|
| Description |
Comparison of gene expression profiles of 20 DPA and 20 DPA cotton fibers from Line 2 and Line 1 respectively
|
| Data processing |
LOWESS normalized, Expression ratios were collected only on those spots with signal intensity >800 in at least one dye channel on the microarray slides.
|
| |
|
| Submission date |
Jul 26, 2011 |
| Last update date |
Jul 22, 2015 |
| Contact name |
Lu You Yuan |
| E-mail(s) |
youluyuan@hotmail.com
|
| Phone |
86-372-2525350
|
| Organization name |
Cotton Research Institute, Chinese Academy of Agricultyral Science
|
| Department |
Molecular Breeding
|
| Lab |
Cotton Lab
|
| Street address |
Huanghe Road 38#
|
| City |
Anyang |
| State/province |
Henan |
| ZIP/Postal code |
455000 |
| Country |
China |
| |
|
| Platform ID |
GPL13969 |
| Series (1) |
| GSE30944 |
cDNA array expression in cotton fiber secondary cell wall development from two germplasm lines that differ in fiber strength |
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