|
Status |
Public on Jul 13, 2011 |
Title |
Untagged Wt_L_ChIP1 |
Sample type |
genomic |
|
|
Channel 1 |
Source name |
Myc ChIP DNA from untagged WT in low iron
|
Organism |
Candida albicans |
Characteristics |
strain: SN250 genotype/variation: wild type chip antibody: Anti-c-Myc 9E10 antibody vendor, cat. number: COVANCE (MMS-150R)
|
Treatment protocol |
Cells were fixed, lysed, immunoprecipitated with anti-Myc antibody and genomic DNAs were sheared by sonication
|
Growth protocol |
Saturated cultures of untagged wild type C. albicans (SN250) was inoculated into iron depleted (YEPD+BPS) liquid medium to OD600=0.05. Cultures were incubated with shaking at 30°C until OD600 0.4.
|
Extracted molecule |
genomic DNA |
Extraction protocol |
Saturated cultures of untagged wild type C. albicans (SN250), Sef1-Myc (SN423), Hap43-Myc (SN840), and Sfu1-Myc (SN646) were inoculated into low iron (untagged, Sef1-Myc, Hap43-Myc) or iron replete (untagged, Sfu1-Myc) liquid medium to OD600=0.05. Cultures were incubated with shaking at 30°C until OD600 0.4, when formaldehyde was added to 1% final (with shaking, room temperature, 15 minutes), followed by glycine to 125mM final (with shaking, room temperature, 5 minutes). Cells were collected by centrifugation at 4°C and washed twice with 20 mM Tris-HCl pH 7.5/150 mM NaCl, followed by freezing in liquid N2 and storage at -80°C. Cell lysis, DNA shearing, and ChIP-Chip were performed as described (Nobile et al., 2009).
|
Label |
Cy5
|
Label protocol |
IP genomic DNA samples were labeled with Cy5 and input genomic DNA samples were labeled with Cy3. Protocol is available in Hernday et al. (Methods Enzymol. 2010;470:737-58)
|
|
|
Channel 2 |
Source name |
Input DNA from untagged WT in low iron
|
Organism |
Candida albicans |
Characteristics |
strain: SN250
|
Treatment protocol |
Cells were fixed, lysed, immunoprecipitated with anti-Myc antibody and genomic DNAs were sheared by sonication
|
Growth protocol |
Saturated cultures of untagged wild type C. albicans (SN250) was inoculated into iron depleted (YEPD+BPS) liquid medium to OD600=0.05. Cultures were incubated with shaking at 30°C until OD600 0.4.
|
Extracted molecule |
genomic DNA |
Extraction protocol |
Saturated cultures of untagged wild type C. albicans (SN250), Sef1-Myc (SN423), Hap43-Myc (SN840), and Sfu1-Myc (SN646) were inoculated into low iron (untagged, Sef1-Myc, Hap43-Myc) or iron replete (untagged, Sfu1-Myc) liquid medium to OD600=0.05. Cultures were incubated with shaking at 30°C until OD600 0.4, when formaldehyde was added to 1% final (with shaking, room temperature, 15 minutes), followed by glycine to 125mM final (with shaking, room temperature, 5 minutes). Cells were collected by centrifugation at 4°C and washed twice with 20 mM Tris-HCl pH 7.5/150 mM NaCl, followed by freezing in liquid N2 and storage at -80°C. Cell lysis, DNA shearing, and ChIP-Chip were performed as described (Nobile et al., 2009).
|
Label |
Cy3
|
Label protocol |
IP genomic DNA samples were labeled with Cy5 and input genomic DNA samples were labeled with Cy3. Protocol is available in Hernday et al. (Methods Enzymol. 2010;470:737-58)
|
|
|
|
Hybridization protocol |
10 independent hybridization experiments were performed on 4 biological replicates of the untagged control and 2 biological replicates each of Sef1-Myc, Sfu1-Myc, and Hap43-Myc. Protocol is available in Hernday et al. (Methods Enzymol. 2010;470:737-58)
|
Scan protocol |
Arrays were scanned on an Axon 4000B scanner per manufacturer's protocol
|
Description |
Control for background
|
Data processing |
Agilent Chip Analytics software v1.2 (Agilent Technologies) was used for initial data normalization and analysis (Tuch et al., 2008), followed by visualization and additional analysis using MochiView v.1.39 (http://johnsonlab.ucsf.edu/). High confidence regulatory events were associated with Agilent segment p-values of ≥ 4 (-log10 p-value based on the enrichment statistic for each probe in the region) and minimum 2-fold (Sef1-Myc and Hap43-Myc) or 1.5-fold (Sfu1-Myc) enrichment in both biological replicates of the epitope-tagged strains (Table S2). Genes with enrichment peaks in untagged controls were excluded.
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|
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Submission date |
Jul 12, 2011 |
Last update date |
Jul 13, 2011 |
Contact name |
Changbin Chen |
E-mail(s) |
Changbin.Chen@ucsf.edu
|
Phone |
4154768992
|
Fax |
4154768201
|
Organization name |
University of California, San Francisco
|
Department |
Department of Microbiology & Immunology
|
Lab |
Dr. Suzanne Noble LAB
|
Street address |
HSE450, 513 Parnassus Avenue
|
City |
San Francisco |
State/province |
California |
ZIP/Postal code |
94143 |
Country |
USA |
|
|
Platform ID |
GPL13696 |
Series (2) |
GSE30591 |
A unique iron homeostasis regulatory circuit with reciprocal roles in Candida albicans commensalism and pathogenesis [ChIP_chip] |
GSE30593 |
A unique iron homeostasis regulatory circuit with reciprocal roles in Candida albicans commensalism and pathogenesis |
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