|
| Status |
Public on Mar 01, 2015 |
| Title |
Mycelium biological sample 2 |
| Sample type |
RNA |
| |
|
| Source name |
Coprinopsis cinerea mycelium
|
| Organism |
Coprinopsis cinerea |
| Characteristics |
developmental stage: mycelium
|
| Treatment protocol |
All sample handling instruments including mortars, pestles, forceps and scissors were prebaked at 300oC overnight to remove RNase and cooled to -20oC before use. Latex gloves and barrier tips were used to ensure minimum RNase contamination.
|
| Growth protocol |
The C. cinerea strain used is a dikaryotic strain (with 50% Okayama-7 origin) backcrossed with Okayama-7, the monokaryotic strain used for the genome sequencing project, for 5 generations. This strain is about 98.44% homologous to Okayama-7 while retaining the ability to fruit. The strain was cultivated on YMD medium containing 0.4% yeast extract, 1% malt extract and 0.4% potato dextrose solidified with Bacto® agar. Mycelium was cultivated on agar plates at 37oC for about 1 week until the mycelium grew over the whole agar surface. Fruiting body development was induced by incubating the mycelia culture at 25oC under a light/dark regime of 12/12 hr. The incubator was kept at a relative humidity higher than 60% for the production of fruiting bodies.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The RNAs were extracted by the RNeasy Plant Mini Kit (Qiagen). Samples were freshly scraped or cut into the mortar soaked with liquid nitrogen and were immediately grinded to fine powder by pestle. About 100mg of samples were transferred to each RNase-free microcentrifuge tube containing 450μl Buffer RLC and the tubes were vortexed to homogenize the samples. The lysate were then transferred to QIAshredder spin column and centrifuged for 2 min at 10000rpm. The supernatant of the flowthrough were transferred to new microcentrifuge tubes. For each tube, 200μl 100% ethanol was added to the cleared lysate and mixed by pipeting. The mixtures together with any precipitates, were transferred to an RNeasy spin column and centrifuged for 15s at 10000rpm. The flow through was discarded. After that, the spin column was washed by 700μl Buffer RW1 followed by 500μl Buffer RPE wash for two times. After drying the spin column by centrifuge for 1 min at full speed, 40μl RNase-free water was added to the column to elute the RNA and kept at -80oC until use .
|
| Label |
Cy3
|
| Label protocol |
The cDNA samples were labeled with Cy3 with the NimbleGen One-color DNA labeling kit according to the manufacturer's protocol (http://www.nimblegen.com/support/dna-microarray-support.htmll).
|
| |
|
| Hybridization protocol |
The labeled cDNAs were hybridized using the NimbleGen mixer and hybridization system (http://www.nimblegen.com/support/dna-microarray-support.htmll).
|
| Scan protocol |
The slides were scanned with NimbleGen MS 200 Microarray Scanner using manufacturer's protocol (http://www.nimblegen.com/support/dna-microarray-support.htmll).
|
| Data processing |
The data were extracted and normalized using the NimbleScan software.
|
| |
|
| Submission date |
Oct 16, 2014 |
| Last update date |
Mar 02, 2015 |
| Contact name |
Xuanjin Cheng |
| E-mail(s) |
xcheng@cuhk.edu.hk
|
| Phone |
852-39431331
|
| Organization name |
The Chinese University of Hong Kong
|
| Department |
School of Life Sciences
|
| Street address |
Rm 386, Science Center South Block
|
| City |
Shatin |
| ZIP/Postal code |
999077 |
| Country |
Hong Kong |
| |
|
| Platform ID |
GPL19301 |
| Series (1) |
| GSE62427 |
A "developmental hourglass" in fungi |
|
