|
| Status |
Public on Dec 28, 2015 |
| Title |
ssn6 (a/a) v. pooled reference (replicate B2) |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
SSN6 homozygous deletion (a/a)
|
| Organism |
Candida albicans |
| Characteristics |
parental strain background: SC5314
source strain: SN87
strain: SSN6 homozygous deletion (a/a)
|
| Growth protocol |
All samples were grown @ 25°C to log phase in synthetic defined (SD) media plus amino acids and uridine; samples labeled replicates A & B were further supplemented with 2.5 mg/ml of arginine to accommodate a growth defect likely associated with the arg- genotype of the WT a/α strain.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Ambion RiboPure Yeast RNA Kit (AM1926)
|
| Label |
cy5
|
| Label protocol |
cDNA was synthesized from either 8 µg (Replicate C) or 6.25 µg (Replicates A&B) total RNA using Superscript II Reverse Transcriptase (Invitrogen, 18064-014), according to the manufacturer’s instructions, and using a mixture of 0.5 mM 3:2 aminoallyl-dUTP (Ambion 8437):dNTPs. A pooled reference was created by taking equal mass proportions of cDNA from each experimental strain, cDNA from the experimental strain was dye coupled to Cy5 (GE Healthcare PA25001). The pooled reference was dye coupled to Cy3 (GE Healthcare PA23001).
|
| |
|
| Channel 2 |
| Source name |
pooled reference
|
| Organism |
Candida albicans |
| Characteristics |
source strains: pooled reference from equal amounts of RNA from each of source strains in ch1
|
| Growth protocol |
All samples were grown @ 25°C to log phase in synthetic defined (SD) media plus amino acids and uridine; samples labeled replicates A & B were further supplemented with 2.5 mg/ml of arginine to accommodate a growth defect likely associated with the arg- genotype of the WT a/α strain.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Ambion RiboPure Yeast RNA Kit (AM1926)
|
| Label |
cy3
|
| Label protocol |
cDNA was synthesized from either 8 µg (Replicate C) or 6.25 µg (Replicates A&B) total RNA using Superscript II Reverse Transcriptase (Invitrogen, 18064-014), according to the manufacturer’s instructions, and using a mixture of 0.5 mM 3:2 aminoallyl-dUTP (Ambion 8437):dNTPs. A pooled reference was created by taking equal mass proportions of cDNA from each experimental strain, cDNA from the experimental strain was dye coupled to Cy5 (GE Healthcare PA25001). The pooled reference was dye coupled to Cy3 (GE Healthcare PA23001).
|
| |
|
| |
| Hybridization protocol |
0.4 µg Cy3-labelled cDNA and 0.4 µg Cy5-labelled cDNA were hybridized onto custom Agilent 8*15k microarrays (AMADID #020166)
|
| Scan protocol |
4000B Axon Instrument Scanner
|
| Description |
ssn6 (a/a)_repB2
|
| Data processing |
Arrays were gridded with Gene Pix Pro software version 7, Lowess normalization was performed using a Goulphar script for R. Following normalization, the 2 or 5 probes per feature were collapsed to their median value prior to further analysis.
|
| |
|
| Submission date |
Oct 14, 2015 |
| Last update date |
Dec 28, 2015 |
| Contact name |
Matthew Lohse |
| Organization name |
BioSynesis, Inc.
|
| Street address |
44 Vicksburg Street
|
| City |
San Francisco |
| State/province |
CA |
| ZIP/Postal code |
94114 |
| Country |
USA |
| |
|
| Platform ID |
GPL13830 |
| Series (1) |
| GSE74011 |
Ssn6 defines a new level of regulation of white-opaque switching in Candida albicans and is required for the stochasticity of the switch |
|
